Dihydropyridin-2(1H)-one compounds as S-nitrosoglutathione reductase inhibitors and neurokinin-3 receptor antagonists

ABSTRACT

The present invention is directed to novel dihydropyridin-2(1H)-one compounds useful as S-nitrosoglutathione reductase (GSNOR) inhibitors and/or Neurokinin-3 (NK3) receptor antagonists, pharmaceutical compositions comprising such compounds, and methods of making and using the same.

RELATED APPLICATIONS

This application is a divisional of U.S. application Ser. No.13/805,626, filed Dec. 19, 2012. U.S. application Ser. No. 13/805,626 isa 35 U.S.C. §371 national phase application of International ApplicationSerial No. PCT/US2011/043374, filed Jul. 8, 2011 (WO 2012/009227).International Application Serial No. PCT/US2011/043374 claims priorityto U.S. Provisional Application Ser. No. 61/365,225, filed Jul. 16,2010. Each of these references is incorporated herein by reference intheir entirety.

FIELD OF THE INVENTION

The present invention is directed to novel dihydropyridin-2(1H)-onecompounds, pharmaceutical compositions comprising such compounds, andmethods of making and using the same. These compounds are useful asinhibitors of S-nitrosoglutathione reductase (GSNOR) and/or antagonistsof the Neurokinin-3 (NK3) receptor.

BACKGROUND OF THE INVENTION

The chemical compound nitric oxide is a gas with chemical formula NO. NOis one of the few gaseous signaling molecules known in biologicalsystems, and plays an important role in controlling various biologicalevents. For example, the endothelium uses NO to signal surroundingsmooth muscle in the walls of arterioles to relax, resulting invasodilation and increased blood flow to hypoxic tissues. NO is alsoinvolved in regulating smooth muscle proliferation, platelet function,neurotransmission, and plays a role in host defense. Although nitricoxide is highly reactive and has a lifetime of a few seconds, it canboth diffuse freely across membranes and bind to many molecular targets.These attributes make NO an ideal signaling molecule capable ofcontrolling biological events between adjacent cells and within cells.NO is a free radical gas, which makes it reactive and unstable, thus NOis short lived in vivo, having a half life of 3-5 seconds underphysiologic conditions.

In the presence of oxygen, NO can combine with thiols to generate abiologically important class of stable NO adducts called S-nitrosothiols(SNO's). This stable pool of NO has been postulated to act as a sourceof bioactive NO and as such appears to be critically important in healthand disease, given the centrality of NO in cellular homeostasis (Stamleret al., Proc. Natl. Acad. Sci. USA, 89:7674-7677 (1992)). Protein SNO'splay broad roles in cardiovascular, respiratory, metabolic,gastrointestinal, immune and central nervous system function (Foster etal., 2003, Trends in Molecular Medicine Volume 9, Issue 4, April 2003,pages 160-168). One of the most studied SNO's in biological systems isS-nitrosoglutathione (GSNO) (Gaston et al., Proc. Natl. Acad. Sci. USA90:10957-10961 (1993)), an emerging key regulator in NO signaling sinceit is an efficient trans-nitrosating agent and appears to maintain anequilibrium with other S-nitrosated proteins (Liu et al., 2001) withincells. Given this pivotal position in the NO—SNO continuum, GSNOprovides a therapeutically promising target to consider when NOmodulation is pharmacologically warranted.

In light of this understanding of GSNO as a key regulator of NOhomeostasis and cellular SNO levels, studies have focused on examiningendogenous production of GSNO and SNO proteins, which occurs downstreamfrom the production of the NO radical by the nitric oxide synthetase(NOS) enzymes. More recently there has been an increasing understandingof enzymatic catabolism of GSNO which has an important role in governingavailable concentrations of GSNO and consequently available NO andSNO's.

Central to this understanding of GSNO catabolism, researchers haverecently identified a highly conserved S-nitrosoglutathione reductase(GSNOR) (Jensen et al., Biochem J., 331:659-668 (1998); Liu et al.,Nature, 410:490-494 (2001)). GSNOR is also known asglutathione-dependent formaldehyde dehydrogenase (GS-FDH), alcoholdehydrogenase 3 (ADH-3) (Uotila and Koivusalo, Coenzymes and Cofactors.,D. Dolphin, ed. pp. 517-551 (New York, John Wiley & Sons, 1989)), andalcohol dehydrogenase 5 (ADH-5). Importantly GSNOR shows greateractivity toward GSNO than other substrates (Jensen et al., 1998; Liu etal., 2001) and appears to mediate important protein and peptidedenitrosating activity in bacteria, plants, and animals. GSNOR appearsto be the major GSNO-metabolizing enzyme in eukaryotes (Liu et al.,2001). Thus, GSNO can accumulate in biological compartments where GSNORactivity is low or absent (e.g. airway lining fluid) (Gaston et al.,1993).

Yeast deficient in GSNOR accumulate S-nitrosylated proteins which arenot substrates of the enzyme, which is strongly suggestive that GSNOexists in equilibrium with SNO-proteins (Liu et al., 2001). Preciseenzymatic control over ambient levels of GSNO and thus SNO-proteinsraises the possibility that GSNO/GSNOR may play roles across a host ofphysiological and pathological functions including protection againstnitrosative stress wherein NO is produced in excess of physiologicneeds. Indeed, GSNO specifically has been implicated in physiologicprocesses ranging from the drive to breathe (Lipton et al., Nature,413:171-174 (2001)) to regulation of the cystic fibrosis transmembraneregulator (Zaman et al., Biochem Biophys Res Commun, 284:65-70 (2001),to regulation of vascular tone, thrombosis and platelet function (deBelder et al., Cardiovasc Res. May; 28(5):691-4. (1994); Z. Kaposzta, Aet al., Circulation; 106(24): 3057-3062, 2002) as well as host defense(de Jesus-Berrios et al., Curr. Biol., 13:1963-1968 (2003)). Otherstudies have found that GSNOR protects yeast cells against nitrosativestress both in vitro (Liu et al., 2001) and in vivo (de Jesus-Berrios etal., 2003).

Collectively data suggest GSNOR as a primary physiological ligand forthe enzyme S-nitrosoglutathione reductase (GSNOR), which catabolizesGSNO and consequently reduces available SNO's and NO in biologicalsystems (Liu et al., 2001), (Liu et al., Cell, (2004), 116(4), 617-628),and (Que et al., Science, 2005, 308, (5728):1618-1621). As such, thisenzyme plays a central role in regulating local and systemic bioactiveNO. Since perturbations in NO bioavailability has been linked to thepathogenesis of numerous disease states, including hypertension,atherosclerosis, thrombosis, asthma, gastrointestinal disorders,inflammation and cancer, agents that regulate GSNOR activity arecandidate therapeutic agents for treating diseases associated withnitric oxide imbalance.

Currently, there is a great need in the art for diagnostics,prophylaxis, ameliorations, and treatments for medical conditionsrelating to increased NO synthesis and/or increased NO bioactivity. Inaddition, there is a significant need for novel compounds, compositionsand methods for preventing, ameliorating, or reversing otherNO-associated disorders. The present invention satisfies these needs.

The mammalian tachykinins, also known as neurokinins, are a family ofsmall peptides that share a common carboxyl-terminal sequence ofPhe-X-Gly-Leu-Met-NH₂ (Maggio et al., Annual Rev. Neuroscience 11:13-28(1998). The main members of the family are substance P (SP), neurokininA (NKA) and neurokinin B (NKB). As neurotransmitters these peptidesexert their biological activity via three distinct neurokinin (NK)receptors termed neurokinin-1 (NK1), neurokinin-2 (NK2) and neurokinin-3(NK3). SP binds preferentially to NK1, NKA to NK2 and NKB to NK3. TheNK3 receptor is characterized by a predominant expression in the centralnervous system (CNS) and its involvement in the modulation of thecentral monoaminergic (noradenaline and dopamine) and amino acid (GABA)neurotransmission. These properties make the NK3 receptor a potentialtarget for CNS diseases such as schizophrenia (Spooren et al., Nat. Rev.Drug Discov. 4:967-975 (2005)).

Schizophrenia is a chronic, severe, and disabling brain disorder thataffects about 1% of the world's population. Symptoms begin in earlyadulthood and are followed by a period of interpersonal and socialdysfunction. The symptoms of schizophrenia fall into three broadcategories: positive symptoms, negative symptoms, and cognitivesymptoms. Positive symptoms include hallucinations, delusions, thoughtdisorders and movement disorders. Negative symptoms include depression,anhedonia, blunted affect, diminished speech and cognitive symptomsinclude memory and attention deficits as well as social withdrawal.

There is no single cause of schizophrenia however, increased dopamineactivity in the mesolimbic pathway of the brain is consistently found inschizophrenic individuals. The lack of knowledge about the exact causeand nature of this disease make development of new drugs difficult.Treatment has been focused on antipsychotic medication which primarilyworks by suppressing dopamine activity. As these drugs have evolvedthrough the years the side effect profile has improved but they stillexhibit some side effects such as weight gain. In 2004 Sanofi-Synthelabopublished clinical results for Osanetant which was identified as apotent and selective antagonist of the NK3 receptor for the treatment ofschizophrenia and in 2005 GSK published clinical results for talnetantwhich was shown to ameliorate the cognitive issues of schizophrenicshowever, both compounds have poor pharmacokinetics and pharmacodynamicproperties including poor solubility, poor bioavailability, relativelyhigh clearance and poor brain-blood barrier penetration. In spite of theliabilities with these compounds, clinical results to date suggest thatthe NK3 receptor may prove to be a promising target for treatment ofschizophrenia providing that pharmacokinetic and pharmacodynamic issuescan be resolved.

Irritable bowel syndrome (IBS) is a chronic, episodic functionalgastrointestinal (GI) disorder characterized by abdominalpain/discomfort and altered bowel habit (constipation, diarrhea oralternating periods of both). Patients often experience additionalsymptoms such as bloating, sensation of incomplete evacuation, straining(constipation) and urgency (diarrhea). IBS patients can experiencesymptoms for many years, with an average duration of 10 or more years.IBS is often unrecognized or untreated, with as few as 25% of IBSsufferers seeking professional health care. IBS prevalence is estimatedto be up to 20% of the population. Functional bowel disorders such asIBS are characterized by visceral hypersensitivity defined by reducedpain and discomfort thresholds, which may manifest as pain associatedwith bowel disturbances (Akbar et al., Alimentary Pharmacology andTherapeutics, 30(5): 423-435 (2009)). Although the pathogenesis ofvisceral hypersensitivity is not fully understood, several mechanismshave been proposed including subtle inflammation, psychosocial factorsand altered sensorimotor function of the gut, a major component of whichis believed to be peripheral and central sensitization of visceralafferent neuronal pathways. Similarly, the other functional boweldisorders such as noncardiac chest pain, functional dyspepsia andfunctional abdominal pain present commonly and treatment of thesedisorders can be challenging. Over the past 30 years, the main treatmentof irritable bowel syndrome has aimed to normalize gastrointestinaltransit using either laxatives or antidiarrheal agents, with or withoutthe concurrent use of spasmolytics. These therapeutic options arelimited and often disappointing in efficacy.

Recent investigation into the pathophysiology of irritable bowelsyndrome has focused on evaluation of visceral hypersensitivity (Buenoet al. Gut, 51 (Suppl):19-23 (2002)). At the same time, more informationhas been acquired on the status of the local immune system as a possiblecause for sensitization of nerve terminals. Such investigations havestimulated the emergence of new concepts and original candidate drugsfor the treatment of this functional disorder.

Tachykinin receptors do not appear to play significant roles in normalGI functions, but may be involved in defensive or pathologicalprocesses. NK3 receptors have been found to mediate certain disruptionsof intestinal motility. The activity may be driven by tachykininsreleased from intrinsic primary afferent neurones (IPANs), which induceslow excitatory postsynaptic potential (EPSP) activity in connectingIPANs and hence, a degree of hypersensitivity within the enteric nervoussystem. The same process is also proposed to increase C-fibresensitivity, either indirectly or directly. Thus, NK3 receptorantagonists inhibit intestinal nociception via a “peripheral” mechanismthat may be intestine-specific. Studies with talnetant and otherselective NK3 receptor antagonists revealed an exciting and novelpathway by which pathological changes in intestinal motility andnociception can be induced, suggesting a role for NK3 receptorantagonism in irritable bowel syndrome (Sanger, Brit. J. of Pharm.,141:1303-1312 (2004)).

Currently, there is a great need in the art for diagnostics,prophylaxis, ameliorations, and treatments for medical conditionsrelating to a disease or condition characterized by overstimulation ofNK3. In addition, there is a significant need for novel compounds,compositions and methods for preventing, ameliorating, or reversingother NK3 associated disorders. The present invention satisfies theseneeds.

SUMMARY OF THE INVENTION

The present invention provides novel dihydropyridin-2(1H)-one compounds.These compounds are useful as S-nitrosoglutathione reductase (“GSNOR”)inhibitors and/or Neurokinin-3 (“NK3”) receptor antagonists. Theinvention encompasses pharmaceutically acceptable salts, prodrugs, andmetabolites of the described compounds. Also encompassed by theinvention are pharmaceutical compositions comprising at least onecompound of the invention and at least one pharmaceutically acceptablecarrier.

The compositions of the present invention can be prepared in anysuitable pharmaceutically acceptable dosage form.

The present invention provides a method for inhibitingS-nitrosoglutathione reductase in a subject in need thereof. Such amethod comprises administering a therapeutically effective amount of apharmaceutical composition comprising at least one GSNOR inhibitor or apharmaceutically acceptable salt thereof, a prodrug or metabolitethereof, in combination with at least one pharmaceutically acceptablecarrier. The GSNOR inhibitor can be a novel compound according to theinvention, or it can be a known compound which previously was not knownto be an inhibitor of GSNOR.

The present invention also provides a method of treating a disorderameliorated by NO donor therapy in a subject in need thereof. Such amethod comprises administering a therapeutically effective amount of apharmaceutical composition comprising at least one GSNOR inhibitor or apharmaceutically acceptable salt thereof, a prodrug, or metabolitethereof, in combination with at least one pharmaceutically acceptablecarrier. The GSNOR inhibitor can be a novel compound according to theinvention, or it can be a known compound which previously was not knownto be an inhibitor of GSNOR.

The present invention also provides a method of treating a cellproliferative disorder in a subject in need thereof. Such a methodcomprises administering a therapeutically effective amount of apharmaceutical composition comprising at least one GSNOR inhibitor or apharmaceutically acceptable salt thereof, a prodrug, or metabolitethereof, in combination with at least one pharmaceutically acceptablecarrier. The GSNOR inhibitor can be a novel compound according to theinvention, or it can be a known compound which previously was not knownto be an inhibitor of GSNOR.

The methods of the invention encompass administration with one or moresecondary active agents. Such administration can be sequential or in acombination composition.

The present invention also provides novel dihydropyridin-2(1H)-onecompounds useful as neurokinin-3 receptor antagonists. The tachykinins,substance P (SP), neurokinin A (NKA), and neurokinin B (NKB), arestructurally similar members of a family of neuropeptides. Each of theseis an agonist of the receptor types, neurokinin-1 receptor (NK1),neurokinin-2 receptor (NK2), and neurokinin-3 receptor (NK3), which areso defined according to their unique amino acid sequence and theirrelative abilities to bind tachykinins with high affinity and to beactivated by the natural agonists, SP, NKA, and NKB, respectively (seealso U.S. Pat. No. 5,434,158, which is herein incorporated byreference). In some embodiments, the dihydropyridin-2(1H)-ones of thepresent invention are NK3 receptor antagonists.

The present invention provides a method for antagonizing theneurokinin-3 (NK3) receptor. Such a method comprises administering atherapeutically effective amount of a pharmaceutical compositioncomprising at least one NK3 receptor antagonist or a pharmaceuticallyacceptable salt thereof, a prodrug or metabolite thereof, in combinationwith at least one pharmaceutically acceptable carrier. The NK3 receptorantagonist can be a novel compound according to the invention, or it canbe a known compound which previously was not known to be an NK3 receptorantagonist.

Although methods and materials similar or equivalent to those describedherein can be used in the practice or testing of the present invention,suitable methods and materials are described below. All publiclyavailable publications, patent applications, patents, and otherreferences mentioned herein are incorporated by reference in theirentirety. In the case of conflict, the present specification, includingdefinitions, will control.

Both the foregoing summary and the following detailed description areexemplary and explanatory and are intended to provide further details ofthe compositions and methods as claimed. Other objects, advantages, andnovel features will be readily apparent to those skilled in the art fromthe following detailed description.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS A. Overview of theInvention

Until recently, S-nitrosoglutathione reductase (GSNOR) was known tooxidize the formaldehyde glutathione adduct, S-hydroxymethylglutathione.GSNOR has since been identified in a variety of bacteria, yeasts, plantsand animals and is well conserved. The proteins from E. coli, S.cerevisiae and mouse macrophages share over 60% amino acid sequenceidentity. GSNOR activity (i.e., decomposition of S-nitrosoglutathionewhen NADH is present as a required cofactor) has been detected in E.coli, in mouse macrophages, in mouse endothelial cells, in mouse smoothmuscle cells, in yeasts, and in human HeLa, epithelial and monocytecells. Human GSNOR nucleotide and amino acid sequence information can beobtained from the National Center for Biotechnology Information (NCBI)databases under Accession Nos. M29872, NM_(—)000671. Mouse GSNORnucleotide and amino acid sequence information can be obtained from NCBIdatabases under Accession Nos. NM_(—)007410. In the nucleotide sequence,the start site and stop site are underlined. CDS designates codingsequence. SNP designates single nucleotide polymorphism. Other relatedGSNOR nucleotide and amino acid sequences, including those of otherspecies, can be found in U.S. Patent Application 2005/0014697.

In accord with the present invention, GSNOR has been shown to functionin vivo and in vitro to metabolize S-nitrosoglutathione (GSNO) andprotein S-nitrosothiols (SNOs) to modulate NO bioactivity, bycontrolling the intracellular levels of low mass NO donor compounds andpreventing protein nitrosylation from reaching toxic levels.

Based on this, it follows that inhibition of this enzyme potentiatesbioactivity in all diseases in which NO donor therapy is indicated,inhibits the proliferation of pathologically proliferating cells, andincreases NO bioactivity in diseases where this is beneficial.

The present invention provides pharmaceutical agents that are potentinhibitors of GSNOR and/or antagonists of the NK3 receptor. Inparticular, provided are substituted dihydropyridin-2(1H)-one analogshaving the structures depicted below (Formula I), or a pharmaceuticallyacceptable salt, stereoisomer, or prodrug thereof.

whereinX is selected from the group consisting of aryl, substituted aryl,heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl,heterocyclyl, and substituted heterocyclyl, each having 6 members orless in the ring;Y is selected from the group consisting of aryl, substituted aryl,heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl,heterocyclyl, and substituted heterocyclyl;Z is selected from the group consisting of O, S, and NR₇;R₁, R₂ and R₇ are independently selected from the group consisting ofhydrogen, and C₁-C₆ alkyl;R₃ is selected from the group consisting of hydrogen, nitro, cyano,carboxyl, carbamoyl, methylsulfonamido, fluoro, chloro, bromo, hydroxy,methylsulfonyl, and methylsulfinyl, isoxazol-4-yl, C₁-C₆ alkoxy,—C(NH)NHOH, sulfonic acid, and acetyl;R₄ is selected from the group consisting of hydrogen, hydroxy, methoxy,carboxyl, and tetrazol-5-yl;wherein, when R₃ is hydrogen, then R₄ is not hydrogen;or optionally R₃ and R₄, taken together can form a heterocycle;R₅ is selected from the group consisting of hydrogen, hydroxyl, carboxy,chloro, fluoro, cyano, —O(CH₂)₁₋₆NMe₂, C₁-C₆ alkyl, —O(CH₂)₁₋₆OCH₃,—O(CH₂)₁₋₆OH, acetyl, CF₃, and C₁-C₆ alkoxy;or optionally R₄ and R₅, taken together can form a heterocycle; andR₆ is selected from the group consisting of hydrogen and hydroxyl.

As used in this context, the term “analog” refers to a compound havingsimilar chemical structure and function as compounds of Formula I thatretains the dihydropyridin-2(1H)-one ring.

Some dihydropyridin-2(1H)-one analogs of the invention can also exist invarious isomeric forms, including configurational, geometric andconformational isomers, as well as existing in various tautomeric forms,particularly those that differ in the point of attachment of a hydrogenatom. As used herein, the term “isomer” is intended to encompass allisomeric forms of a compound including tautomeric forms of the compound.

Illustrative compounds having asymmetric centers can exist in differentenantiomeric and diastereomeric forms. A compound can exist in the formof an optical isomer or a diastereomer. Accordingly, the inventionencompasses compounds in the forms of their optical isomers,diastereomers and mixtures thereof, including racemic mixtures.

It should be noted that if there is a discrepancy between a depictedstructure and a name given to that structure, the depicted structurecontrols. In addition, if the stereochemistry of a structure or aportion of a structure is not indicated with, for example, bold, wedged,or dashed lines, the structure or portion of the structure is to beinterpreted as encompassing all stereoisomers of the described compound.

B. S-Nitrosoglutathione Reductase Inhibitors and/or NK3 ReceptorAntagonists

1. Inventive Compounds

In one of its aspects the present invention provides a compound having astructure shown in Formula I, or a pharmaceutically acceptable salt,stereoisomer, or prodrug thereof:

whereinX is selected from the group consisting of aryl, substituted aryl,heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl,heterocyclyl, and substituted heterocyclyl, each having 6 members orless in the ring;Y is selected from the group consisting of aryl, substituted aryl,heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl,heterocyclyl, and substituted heterocyclyl;Z is selected from the group consisting of O, S, and NR₇;R₁, R₂ and R₇ are independently selected from the group consisting ofhydrogen, and C₁-C₆ alkyl;R₃ is selected from the group consisting of hydrogen, nitro, cyano,carboxyl, carbamoyl, methylsulfonamido, fluoro, chloro, bromo, hydroxyl,methylsulfonyl, and methylsulfinyl, isoxazol-4-yl, C₁-C₆ alkoxy,—C(NH)NHOH, sulfonic acid, and acetyl;R₄ is selected from the group consisting of hydrogen, hydroxyl, methoxy,carboxyl, and tetrazol-5-yl;wherein, when R₃, is hydrogen, then R₄ is not hydrogen;or optionally R₃ and R₄, taken together can form a heterocycle; andR₅ is selected from the group consisting of hydrogen, hydroxyl,carboxyl, chloro, fluoro, cyano, —O(CH₂)₁₋₆NMe₂, C₁-C₆ alkyl,—O(CH₂)₁₋₆OCH₃, —O(CH₂)₁₋₆OH, acetyl, CF₃, and C₁-C₆ alkoxy;or optionally R₄ and R₅, taken together can form a heterocycle; andR₆ is selected from the group consisting of hydrogen and hydroxyl.

In a further aspect of the invention, R4 is selected from the groupconsisting of hydroxyl, carboxyl, and tetrazol-5-yl.

In a further aspect of the invention, R1, R2 and R7 are independentlyselected from the group consisting of hydrogen and methyl;

R₃ is selected from the group consisting of hydrogen, nitro, cyano,carboxyl, carbamoyl, methylsulfonamido, fluoro, chloro, bromo, hydroxyl,methylsulfonyl, and methylsulfinyl, isoxazol-4-yl, C₁-C₆ alkoxy,—C(NH)NHOH, sulfonic acid, acetyl;

R₄ is selected from the group consisting of hydroxyl, carboxyl, andtetrazol-5-yl;

R₅ is selected from the group consisting of hydrogen, hydroxyl, carboxy,chloro, fluoro, cyano, —O(CH₂)₂NMe₂, C₁-C₆ alkyl, —O(CH₂)₂OCH₃,—O(CH₂)₂OH, acetyl, CF₃, methoxy, ethoxy, isopropoxy, and n-propoxy; and

R₆ is hydrogen.

In a further aspect of the invention, R₃ is selected from the groupconsisting of hydrogen, nitro, and hydroxyl; R₄ is selected from thegroup consisting of hydroxyl, carboxyl, and tetrazol-5-yl; and R₅ isselected from the group consisting of hydrogen, ethoxy, fluoro, and—O(CH₂)₂OH.

In a further aspect of the invention, suitable identities for X include,but are not limited to aryl, substituted aryl, heteroaryl, andsubstituted heteroaryl.

In a further aspect of the invention, suitable identities for X include,but are not limited to phenyl, substituted phenyl, thiophen-yl,substituted thiophen-yl, thiazol-yl, substituted thiazol-yl, pyrazin-yl,substituted pyrazin-yl, pyridin-yl, and substituted pyridin-yl,cyclohexyl, substituted cyclohexyl.

In a further aspect of the invention, suitable identities for X include,but are not limited to, phenyl, thiophen-2-yl, thiophen-3-yl,thiazol-2-yl, 2-fluorophenyl, p-tolyl, m-tolyl, biphenyl-4-yl,4-methoxyphenyl, 3-chlorophenyl, 3,4-dichlorophenyl, 3-methoxyphenyl,3,4-dimethoxyphenyl, 4-bromophenyl, o-tolyl, 4-chlorophenyl,2-chlorophenyl, 3-cyanophenyl, 3,4-difluorophenyl, 4-cyanophenyl,3-carbamoylphenyl, pyrazin-2-yl, biphenyl-3-yl, 2-cyanophenyl,pyridin-4-yl, and pyridin-3-yl, 4-(dimethylamino)phenyl, 3-fluorophenyl,3-ethylphenyl, and cyclohexyl.

In a further aspect of the invention, suitable identities for X include,but are not limited to, phenyl, thiophen-2-yl, thiophen-3-yl, andpyridin-3-yl.

In a further aspect of the invention, suitable identities for Y include,but are not limited to aryl, substituted aryl, heteroaryl, substitutedaryl, cycloalkyl, and substituted cycloalkyl.

In a further aspect of the invention, suitable identities for Y include,but are not limited to phenyl, substituted phenyl, thiophen-yl,substituted thiophen-yl, thiazol-yl, substituted thiazol-yl, pyrazin-yl,substituted pyrazin-yl, pyridin-yl, substituted pyridin-yl, furan-yl,substituted furan-yl, benzo[d][1,3]dioxol-yl, substitutedbenzo[d][1,3]dioxol-yl, imidazol-yl, substituted imidazol-yl,naphthalen-yl, substituted naphthalen-yl, pyrrol-yl, substitutedpyrrol-yl, pyrazol-yl, substituted pyrazol-yl, tetrahydrofuran-yl,substituted tetrahydrofuran-yl, cyclopentyl, substituted cyclopentyl,cyclohexyl, and substituted cyclohexyl.

In a further aspect of the invention, suitable identities for Y include,but are not limited to, phenyl, 3-methoxyphenyl, p-tolyl,4-methoxyphenyl, 3,5-dichlorophenyl, 3-fluorophenyl, 4-bromophenyl,biphenyl-4-yl, 4-fluorophenyl, 4-chlorophenyl, 3-chlorophenyl,3,4-dimethoxyphenyl, 3-fluoro-4-methoxyphenyl, 4-chloro-3-fluorophenyl,3-chloro-4-fluorophenyl, 3,4-difluorophenyl, 3,5-difluorophenyl,3,4-dichlorophenyl, 4-hydroxyphenyl, 2,4-difluorophenyl, furan-3-yl,2-chlorophenyl, 3-cyanophenyl, 4-(dimethylamino)phenyl, 2-fluorophenyl,4-morpholinophenyl, 4-aminophenyl, pyridin-2-yl,benzo[d][1,3]dioxol-5-yl, 4-cyanophenyl, pyridin-3-yl, pyridin-4-yl,4-acetamidophenyl, thiophen-2-yl, thiophen-3-yl,1-methyl-1H-imidazol-4-yl, naphthalen-1-yl, methyl phenylcarbamate, andnaphthalen-2-yl, 4-(methanesulfonamido)phenyl, 1H-pyrrol-3-yl,1-(phenylsulfonyl)-1H-pyrrol-3-yl, furan-2-yl,4-(trifluoromethyl)phenyl, o-tolyl, 1-methyl-1H-pyrazol-4-yl,1-methyl-1H-pyrazol-3-yl, 3-chloro-5-fluorophenyl, 3-hydroxyphenyl,pyrazin-2-yl, quinolin-6-yl, isoquinolin-6-yl, 1-methyl-1H-pyrazol-5-yl,tetrahydrofuran-2-yl, cyclopentyl, tetrahydrofuran-3-yl, and cyclohexyl.

In a further aspect of the invention, suitable identities for Y include,but are not limited to phenyl, pyridin-3-yl, 1-methyl-1H-pyrazol-4-yl,and cyclohexyl.

In a further aspect of the invention, Z is O.

In a further aspect of the invention, suitable compounds of formula Iinclude, but are not limited to:

-   4-(3-ethoxy-4-hydroxy-5-nitrophenyl)-5,6-diphenyl-3,4-dihydropyridin-2(1H)-one;-   4-(3-ethoxy-4-hydroxy-5-nitrophenyl)-6-(pyridin-3-yl)-5-(thiophen-3-yl)-3,4-dihydropyridin-2(1H)-one;-   4-(4-(2H-tetrazol-5-yl)phenyl)-5-phenyl-6-(pyridin-3-yl)-3,4-dihydropyridin-2(1H)-one;-   4-(3-ethoxy-4-hydroxy-5-nitrophenyl)-6-(1-methyl-1H-pyrazol-4-yl)-5-(thiophen-3-yl)-3,4-dihydropyridin-2(1H)-one;-   4-(4-(2H-tetrazol-5-yl)phenyl)-6-(1-methyl-1H-pyrazol-4-yl)-5-(thiophen-3-yl)-3,4-dihydropyridin-2(1H)-one;-   2-hydroxy-4-(2-oxo-5,6-diphenyl-1,2,3,4-tetrahydropyridin-4-yl)benzoic    acid;-   2-hydroxy-4-(2-oxo-6-phenyl-5-(thiophen-3-yl)-1,2,3,4-tetrahydropyridin-4-yl)benzoic    acid;-   4-(4-(2H-tetrazol-5-yl)phenyl)-6-(1-methyl-1H-pyrazol-4-yl)-5-phenyl-3,4-dihydropyridin-2(1H)-one;-   2-hydroxy-4-(6-(1-methyl-1H-pyrazol-4-yl)-2-oxo-5-(thiophen-3-yl)-1,2,3,4-tetrahydropyridin-4-yl)benzoic    acid;-   2-fluoro-6-hydroxy-4-(2-oxo-5,6-diphenyl-1,2,3,4-tetrahydropyridin-4-yl)benzoic    acid;-   2-fluoro-6-hydroxy-4-(6-(1-methyl-1H-pyrazol-4-yl)-2-oxo-5-(thiophen-3-yl)-1,2,3,4-tetrahydropyridin-4-yl)benzoic    acid;-   2-ethoxy-6-hydroxy-4-(2-oxo-5,6-diphenyl-1,2,3,4-tetrahydropyridin-4-yl)benzoic    acid;-   4-(6-cyclohexyl-2-oxo-5-(thiophen-3-yl)-1,2,3,4-tetrahydropyridin-4-yl)-2-fluoro-6-hydroxybenzoic    acid;-   4-(4-hydroxy-3-(2-hydroxyethoxy)-5-nitrophenyl)-5,6-diphenyl-3,4-dihydropyridin-2(1H)-one;-   2-fluoro-6-hydroxy-4-(1-methyl-2-oxo-5,6-diphenyl-1,2,3,4-tetrahydropyridin-4-yl)benzoic    acid;-   4-(3-ethoxy-4-hydroxy-5-nitrophenyl)-1-methyl-5,6-diphenyl-3,4-dihydropyridin-2(1H)-one;    and-   2-fluoro-6-hydroxy-4-(6-(1-methyl-1H-pyrazol-4-yl)-2-oxo-5-(thiophen-2-yl)-1,2,3,4-tetrahydropyridin-4-yl)benzoic    acid.

When a bond to a substituent is shown to cross a bond connecting twoatoms in a ring, then such substituent may be bonded to any atom in thering. When a substituent is listed without indicating the atom via whichsuch substituent is bonded to the rest of the compound of a givenformula, then such substituent may be bonded via any atom in suchsubstituent. Combinations of substituents and/or variables arepermissible, but only if such combinations result in stable compounds.

The compounds described herein may have asymmetric centers. Compounds ofthe present invention containing an asymmetrically substituted atom maybe isolated in optically active or racemic forms. It is well known inthe art how to prepare optically active forms, such as by resolution ofracemic forms or by synthesis from optically active starting materials.Many geometric isomers of olefins, C═N double bonds, and the like canalso be present in the compounds described herein, and all such stableisomers are contemplated in the present invention. Cis and transgeometric isomers of the compounds of the present invention aredescribed and may be isolated as a mixture of isomers or as separatedisomeric forms. All chiral, diastereomeric, racemic, and geometricisomeric forms of a structure are intended, unless the specificstereochemistry or isomeric form is specifically indicated. Alltautomers of shown or described compounds are also considered to be partof the present invention.

It is to be understood that isomers arising from such asymmetry (e.g.,all enantiomers and diastereomers) are included within the scope of theinvention, unless indicated otherwise. Such isomers can be obtained insubstantially pure form by classical separation techniques and bystereochemically controlled synthesis. Furthermore, the structures andother compounds and moieties discussed in this application also includeall tautomers thereof. Alkenes can include either the E- or Z-geometry,where appropriate.

2. Representative Compounds

Examples 1-17 list representative novel dihydropyridin-2(1H)-one analogsof Formula I. The synthetic methods that can be used to prepare eachcompound are detailed in Examples 1-17, with reference to intermediatesdescribed in Example 18. Supporting mass spectrometry data and protonNMR data for each compound is also included in Examples 1-17.

GSNOR inhibitor activity was determined by the assay described inExample 19 and IC₅₀ values were obtained. GSNOR inhibitor compounds inExamples 1-17 had an IC₅₀ of about <1.0 μM. GSNOR inhibitor compounds inExamples 1-2, 4, 6-7, 9-14, and 17 had an IC₅₀ of about <0.1 μM.

NK3 receptor antagonist activity was determined for a subset of the 17compounds. A percent inhibition value at the concentration of 10 μM wasdetermined for selected compounds in the human NK3 receptor bindingassay described in Example 20. Compounds in the following Examples hadabout equal to or greater than 50% inhibition at 10 μM: Examples 1-2,and 4. A percent inhibition value at the concentration of 1 μM wasdetermined for selected compounds in the human NK3 receptor bindingassay described in Example 21. The following compounds had about equalto or greater than 50% inhibition at 1 μM: Examples 14 and 16. The humanNK3 receptor binding assay described in Example 22 is used to determineIC₅₀ values. An IC₅₀ of about 1.1 μM was obtained for the compound inExample 1.

In certain embodiments of the invention it has been demonstrated thatracemic mixtures have GSNOR inhibitor activity. Without being bound bytheory, it is believed that when the enantiomers are separated, one ofthe enantiomers has the majority of the GSNOR inhibitor activity and theother enantiomer is significantly less active as a GSNOR inhibitor.Without being bound by theory, it is believed that when the enantiomersof a GSNOR inhibitor are separated, the enantiomer which demonstratessignificantly better GSNOR inhibitor activity is of the S configuration.It has been shown that a structurally related compound4-(3-ethoxy-4-hydroxy-5-nitrophenyl)-5,6-diphenyl-3,4-dihydropyrimidin-2(1H)-onehas an S configuration by X-ray crystallography when the activeenantiomer for GSNOR was crystallized with GSNOR (see PCT applicationUS2010/050164 and PCT application US2010/050186 incorporated herein byreference in their entirety). The S configuration of that compound hasan IC₅₀ of 11 nM as a GSNOR inhibitor, and an IC₅₀ of 19000 nM as an NK3receptor antagonist. The other enantiomer, the R configuration, has anIC₅₀ of 19720 nM as a GSNOR inhibitor, and an IC₅₀ of 110 nM as an NK3receptor antagonist.

In certain embodiments of the invention it has been demonstrated thatracemic mixtures have NK3 receptor antagonist activity as well as GSNORactivity. Without being bound by theory, it is believed that when theenantiomers are separated, one of the enantiomers has the majority ofthe NK3 receptor antagonist activity and the other enantiomer issignificantly less active as a NK3 receptor antagonist. Without beingbound by theory, it is believed that the enantiomer having substantiallyreduced GSNOR inhibitor activity has significant activity for theneurokinin receptor NK3 and that the enantiomer which demonstratessignificantly better NK3 receptor antagonist activity is of the Rconfiguration.

C. Definitions

As used herein, “about” will be understood by persons of ordinary skillin the art and will vary to some extent on the context in which it isused. If there are uses of the term which are not clear to persons ofordinary skill in the art given the context in which it is used, “about”will mean up to plus or minus 10% of the particular term.

The term “acyl” includes compounds and moieties that contain the acetylradical (CH₃CO—) or a carbonyl group to which a straight or branchedchain lower alkyl residue is attached.

The term “alkyl” as used herein refers to a straight or branched chain,saturated hydrocarbon having the indicated number of carbon atoms. Forexample, (C₁-C₆) alkyl is meant to include, but is not limited tomethyl, ethyl, propyl, isopropyl, butyl, sec-butyl, tert-butyl, pentyl,isopentyl, neopentyl, hexyl, isohexyl, and neohexyl. An alkyl group canbe unsubstituted or optionally substituted with one or more substituentsas described herein.

The term “alkenyl” as used herein refers to a straight or branched chainunsaturated hydrocarbon having the indicated number of carbon atoms andat least one double bond. Examples of a (C₂-C₈) alkenyl group include,but are not limited to, ethylene, propylene, 1-butylene, 2-butylene,isobutylene, sec-butylene, 1-pentene, 2-pentene, isopentene, 1-hexene,2-hexene, 3-hexene, isohexene, 1-heptene, 2-heptene, 3-heptene,isoheptene, 1-octene, 2-octene, 3-octene, 4-octene, and isooctene. Analkenyl group can be unsubstituted or optionally substituted with one ormore substituents as described herein.

The term “alkynyl” as used herein refers to a straight or branched chainunsaturated hydrocarbon having the indicated number of carbon atoms andat least one triple bond. Examples of a (C₂-C₈) alkynyl group include,but are not limited to, acetylene, propyne, 1-butyne, 2-butyne,1-pentyne, 2-pentyne, 1-hexyne, 2-hexyne, 3-hexyne, 1-heptyne,2-heptyne, 3-heptyne, 1-octyne, 2-octyne, 3-octyne and 4-octyne. Analkynyl group can be unsubstituted or optionally substituted with one ormore substituents as described herein.

The term “alkoxy” as used herein refers to an —O-alkyl group having theindicated number of carbon atoms. For example, a (C₁-C₆) alkoxy groupincludes —O-methyl, —O-ethyl, —O-propyl, —O-isopropyl, —O-butyl,—O-sec-butyl, —O-tert-butyl, —O-pentyl, —O— isopentyl, —O-neopentyl,—O-hexyl, —O-isohexyl, and —O-neohexyl.

The term “aminoalkyl” as used herein, refers to an alkyl group(typically one to six carbon atoms) wherein one or more of the C₁-C₆alkyl group's hydrogen atoms is replaced with an amine of formula—N(R^(c))₂, wherein each occurrence of R^(c) is independently —H or(C₁-C₆) alkyl. Examples of aminoalkyl groups include, but are notlimited to, —CH₂NH₂, —CH₂CH₂NH₂, —CH₂CH₂CH₂NH₂, —CH₂CH₂CH₂CH₂NH₂,—CH₂CH₂CH₂CH₂CH₂NH₂, —CH₂CH₂CH₂CH₂CH₂CH₂NH₂, —CH₂CH₂CH₂N(CH₃)₂,t-butylaminomethyl, isopropylaminomethyl and the like.

The term “aryl” as used herein refers to a 5- to 14-membered monocyclic,bicyclic or tricyclic aromatic ring system. Examples of an aryl groupinclude phenyl and naphthyl. An aryl group can be unsubstituted oroptionally substituted with one or more substituents as described hereinbelow. Examples of aryl groups include phenyl or aryl heterocycles suchas, pyrrole, furan, thiophene, thiazole, isothiazole, imidazole,triazole, tetrazole, pyrazole, oxazole, isoxazole, pyridine, pyrazine,pyridazine, and pyrimidine, and the like.

As used herein, the term “bioactivity” indicates an effect on one ormore cellular or extracellular process (e.g., via binding, signaling,etc.) which can impact physiological or pathophysiological processes.

The term “carbonyl” includes compounds and moieties which contain acarbon connected with a double bond to an oxygen atom. Examples ofmoieties containing a carbonyl include, but are not limited to,aldehydes, ketones, carboxylic acids, amides, esters, anhydrides, etc.

The term “carboxy” or “carboxyl” means a —COOH group or carboxylic acid.

The term “C_(m)-C_(n)” means “m” number of carbon atoms to “n” number ofcarbon atoms. For example, the term “C₁-C₆” means one to six carbonatoms (C₁, C₂, C₃, C₄, C₅ or C₆). The term “C₂-C₆” includes two to sixcarbon atoms (C₂, C₃, C₄, C₅ or C₆). The term “C₃-C₆” includes three tosix carbon atoms (C₃, C₄, C₅ or C₆).

The term “cycloalkyl” as used herein refers to a 3- to 14-memberedsaturated or unsaturated non-aromatic monocyclic, bicyclic or tricyclichydrocarbon ring system. Included in this class are cycloalkyl groupswhich are fused to a benzene ring. Representative cycloalkyl groupsinclude, but are not limited to, cyclopropyl, cyclobutyl, cyclobutenyl,cyclopentyl, cyclopentenyl, cyclopentadienyl, cyclohexyl, cyclohexenyl,1,3-cyclohexadienyl, cycloheptyl, cycloheptenyl, 1,3-cycloheptadienyl,1,4-cycloheptadienyl, -1,3,5-cycloheptatrienyl, cyclooctyl,cyclooctenyl, 1,3-cyclooctadienyl, 1,4-cyclooctadienyl,-1,3,5-cyclooctatrienyl, decahydronaphthalene, octahydronaphthalene,hexahydronaphthalene, octahydroindene, hexahydroindene, tetrahydroinden,decahydrobenzocycloheptene, octahydrobenzocycloheptene,hexahydrobenzocycloheptene, tetrahydrobenzocyclopheptene,dodecahydroheptalene, decahydroheptalene, octahydroheptalene,hexahydroheptalene, and tetrahydroheptalene,(1s,3s)-bicyclo[1.1.0]butane, bicyclo[1.1.1]pentane,bicyclo[2.1.1]hexane, Bicyclo[2.2.1]heptane, bicyclo[2.2.2]octane,bicyclo[3.1.1]heptane, bicyclo[3.2.1]octane, bicyclo[3.3.1]nonane,bicyclo[3.3.2]decane, bicyclo[3.3.]undecane, bicyclo[4.2.2]decane,bicyclo[4.3.1]decane. A cycloalkyl group can be unsubstituted oroptionally substituted with one or more substituents as described hereinbelow.

The term “halogen” includes fluorine, bromine, chlorine, iodine, etc.

The term “haloalkyl,” as used herein, refers to a C₁-C₆ alkyl groupwherein from one or more of the C₁-C₆ alkyl group's hydrogen atom isreplaced with a halogen atom, which can be the same or different.Examples of haloalkyl groups include, but are not limited to,trifluoromethyl, 2,2,2-trifluoroethyl, 4-chlorobutyl, 3-bromopropyl,pentachloroethyl, and 1,1,1-trifluoro-2-bromo-2-chloroethyl.

The term “heteroalkyl,” by itself or in combination with another term,means, unless otherwise stated, a stable straight or branched chainalkyl, or combinations thereof, consisting of carbon atoms and from oneto three heteroatoms selected from the group consisting of O, N and S,and wherein the nitrogen and sulfur atoms may optionally be oxidized andthe nitrogen heteroatom may optionally be quaternized. The heteroatom(s)O, N and S can be placed at any position of the heteroalkyl group.Examples include —CH₂—CH₂—O—CH₃, —CH₂—CH₂—NH—CH₃, —CH₂—CH₂—N(CH₃)—CH₃,—CH₂—S—CH₂—CH₃, —CH₂—CH₂—S(O)—CH₃, —CH₂—CH₂—S(O)₂—CH₃, and—CH₂—CH═N—OCH₃. Up to two heteroatoms can be consecutive, such as, forexample, —CH₂—NH—OCH₃. When a prefix such as (C₂-C₈) is used to refer toa heteroalkyl group, the number of carbons (2 to 8, in this example) ismeant to include the heteroatoms as well. For example, a C₂-heteroalkylgroup is meant to include, for example, —CH₂OH (one carbon atom and oneheteroatom replacing a carbon atom) and —CH₂SH.

To further illustrate the definition of a heteroalkyl group, where theheteroatom is oxygen, a heteroalkyl group can be an oxyalkyl group. Forinstance, (C₂-C₅) oxyalkyl is meant to include, for example —CH₂—O—CH₃(a C₃-oxyalkyl group with two carbon atoms and one oxygen replacing acarbon atom), —CH₂CH₂CH₂CH₂OH, —OCH₂CH₂OCH₂CH₂OH, —OCH₂CH(OH)CH₂OH, andthe like.

The term “heteroaryl” as used herein refers to an aromatic heterocyclering of 5 to 14 members and having at least one heteroatom selected fromnitrogen, oxygen and sulfur, and containing at least 1 carbon atom,including monocyclic, bicyclic, and tricyclic ring systems.Representative heteroaryls are triazolyl, tetrazolyl, oxadiazolyl,pyridyl, furyl, benzofuranyl, thienyl, benzothienyl, quinolinyl,pyrrolyl, indolyl, oxazolyl, benzoxazolyl, imidazolyl, benzimidazolyl,thiazolyl, benzothiazolyl, isoxazolyl, pyrazolyl, isothiazolyl,pyridazinyl, pyrimidinyl, pyrazinyl, triazinyl, cinnolinyl,phthalazinyl, quinazolinyl, pyrimidyl, azepinyl, oxepinyl, quinoxalinyland oxazolyl. A heteroaryl group can be unsubstituted or optionallysubstituted with one or more substituents as described herein below.

As used herein, the term “heteroatom” is meant to include oxygen (O),nitrogen (N), and sulfur (S).

As used herein, the term “heterocycle” refers to 3- to 14-membered ringsystems which are either saturated, unsaturated, or aromatic, and whichcontains from 1 to 4 heteroatoms independently selected from nitrogen,oxygen and sulfur, and wherein the nitrogen and sulfur heteroatoms canbe optionally oxidized, and the nitrogen heteroatom can be optionallyquaternized, including, including monocyclic, bicyclic, and tricyclicring systems. The bicyclic and tricyclic ring systems may encompass aheterocycle or heteroaryl fused to a benzene ring. The heterocycle canbe attached via any heteroatom or carbon atom, where chemicallyacceptable. Heterocycles include heteroaryls as defined above.Representative examples of heterocycles include, but are not limited to,aziridinyl, oxiranyl, thiiranyl, triazolyl, tetrazolyl, azirinyl,diaziridinyl, diazirinyl, oxaziridinyl, azetidinyl, azetidinonyl,oxetanyl, thietanyl, piperidinyl, piperazinyl, morpholinyl, pyrrolyl,oxazinyl, thiazinyl, diazinyl, dioxanyl, triazinyl, tetrazinyl,imidazolyl, tetrazolyl, pyrrolidinyl, isoxazolyl, furanyl, furazanyl,pyridinyl, oxazolyl, benzoxazolyl, benzisoxazolyl, thiazolyl,benzthiazolyl, thienyl, pyrazolyl, triazolyl, pyrimidinyl,benzimidazolyl, isoindolyl, indazolyl, benzodiazolyl, benzotriazolyl,benzoxazolyl, benzisoxazolyl, purinyl, indolyl, isoquinolinyl,quinolinyl and quinazolinyl. A heterocycle group can be unsubstituted oroptionally substituted with one or more substituents as described hereinbelow.

The term “heterocycloalkyl,” by itself or in combination with otherterms, represents, unless otherwise stated, cyclic versions of“heteroalkyl.” Additionally, a heteroatom can occupy the position atwhich the heterocycle is attached to the remainder of the molecule.Examples of heterocycloalkyl include 1-(1,2,5,6-tetrahydropyridyl),1-piperidinyl, 2-piperidinyl, 3-piperidinyl, 4-morpholinyl,3-morpholinyl, tetrahydrofuran-2-yl, tetrahydrofuran-3-yl,tetrahydrothien-2-yl, tetrahydrothien-3-yl, 1-piperazinyl,2-piperazinyl, and the like.

The term “hydroxyalkyl,” as used herein, refers to an alkyl group havingthe indicated number of carbon atoms wherein one or more of the hydrogenatoms in the alkyl group is replaced with an —OH group. Examples ofhydroxyalkyl groups include, but are not limited to, —CH₂OH, —CH₂CH₂OH,—CH₂CH₂CH₂OH, —CH₂CH₂CH₂CH₂OH, —CH₂CH₂CH₂CH₂CH₂OH,—CH₂CH₂CH₂CH₂CH₂CH₂OH, and branched versions thereof.

The term “hydroxy” or “hydroxyl” includes groups with an —OH or —O⁻.

As used herein and unless otherwise indicated, the term “stereoisomer”means one stereoisomer of a compound that is substantially free of otherstereoisomers of that compound. For example, a stereomerically purecompound having one chiral center will be substantially free of theopposite enantiomer of the compound. A stereomerically pure compoundhaving two chiral centers will be substantially free of otherdiastereomers of the compound. In some embodiments, a stereomericallypure compound comprises greater than about 80% by weight of onestereoisomer of the compound and less than about 20% by weight of otherstereoisomers of the compound, for example greater than about 90% byweight of one stereoisomer of the compound and less than about 10% byweight of the other stereoisomers of the compound, or greater than about95% by weight of one stereoisomer of the compound and less than about 5%by weight of the other stereoisomers of the compound, or greater thanabout 97% by weight of one stereoisomer of the compound and less thanabout 3% by weight of the other stereoisomers of the compound.

As used herein, “protein” is used synonymously with “peptide,”“polypeptide,” or “peptide fragment”. A “purified” polypeptide, protein,peptide, or peptide fragment is substantially free of cellular materialor other contaminating proteins from the cell, tissue, or cell-freesource from which the amino acid sequence is obtained, or substantiallyfree from chemical precursors or other chemicals when chemicallysynthesized.

As used herein, “modulate” is meant to refer to an increase or decreasein the levels of a peptide or a polypeptide, or to increase or decreasethe stability or activity of a peptide or a polypeptide. The term“inhibit” is meant to refer to a decrease in the levels of a peptide ora polypeptide or to decrease in the stability or activity of a peptideor a polypeptide. In preferred embodiments, the peptide which ismodulated or inhibited is S-nitrosoglutathione (GSNO) or proteinS-nitrosothiols (SNOs).

As used here, the terms “nitric oxide” and “NO” encompass unchargednitric oxide and charged nitric oxide species, particularly includingnitrosonium ion (NO⁺) and nitroxyl ion (NO⁻). The reactive form ofnitric oxide can be provided by gaseous nitric oxide. Compounds havingthe structure X—NO_(y) wherein X is a nitric oxide releasing, deliveringor transferring moiety, including any and all such compounds whichprovide nitric oxide to its intended site of action in a form active fortheir intended purpose, and Y is 1 or 2.

As utilized herein, the term “pharmaceutically acceptable” meansapproved by aregulatory agency of a federal or a state government orlisted in the U.S. Pharmacopoeia or other generally recognizedpharmacopoeia for use in animals and, more particularly, in humans. Theterm “carrier” refers to a diluent, adjuvant, excipient, or vehicle withwhich the therapeutic is administered and includes, but is not limitedto such sterile liquids as water and oils.

A “pharmaceutically acceptable salt” or “salt” of a compound of theinvention is a product of the disclosed compound that contains an ionicbond, and is typically produced by reacting the disclosed compound witheither an acid or a base, suitable for administering to a subject. Apharmaceutically acceptable salt can include, but is not limited to,acid addition salts including hydrochlorides, hydrobromides, phosphates,sulphates, hydrogen sulphates, alkylsulphonates, arylsulphonates,arylalkylsulfonates, acetates, benzoates, citrates, maleates, fumarates,succinates, lactates, and tartrates; alkali metal cations such as Li,Na, K, alkali earth metal salts such as Mg or Ca, or organic aminesalts.

A “pharmaceutical composition” is a formulation comprising the disclosedcompounds in a form suitable for administration to a subject. Apharmaceutical composition of the invention is preferably formulated tobe compatible with its intended route of administration. Examples ofroutes of administration include, but are not limited to, oral andparenteral, e.g., intravenous, intradermal, subcutaneous, inhalation,topical, transdermal, transmucosal, and rectal administration.

The term “substituted,” as used herein, means that any one or morehydrogens on the designated atom is replaced with a selection from theindicated group, provided that the designated atom's normal valency isnot exceeded, and that the substitution results in a stable compound.When a substituent is keto (i.e., ═O), then 2 hydrogens on the atom arereplaced. Ring double bonds, as used herein, are double bonds that areformed between two adjacent ring atoms (e.g., C═C, C═N, or N═N).

Substituents for the groups referred to as alkyl, heteroalkyl, alkylene,alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, cycloalkenyl andheterocycloalkenyl can be selected from a variety of groups including—OR^(d)′, ═O, ═NR^(d)′, ═N—OR^(d)′, —NR^(d)′R^(d)″, —SR^(d)′, -halo,—SiR^(d)′R^(d)″R^(d)′″, —OC(O)R^(d)′, —C(O)R^(d)′, —CO₂R^(d)′,—CONR^(d)′R^(d)″, —OC(O)NR^(d)′R^(d)″, —NR^(d)″C(O)R^(d)′,—NR^(d)′″C(O)NR^(d)′R^(d)″, —NR^(d)′″SO₂NR^(d)′R^(d)″,—NR^(d)″CO₂R^(d)′, —NHC(NH₂)═NH, —NR^(a)′C(NH₂)═NH, —NHC(NH₂)═NR^(d)′,—S(O)R^(d)′, —SO₂R^(d)′, —SO₂NR^(d)′R^(d)″, —NR^(d)″SO₂R^(d)′, —CN and—NO₂, in a number ranging from zero to three, with those groups havingzero, one or two substituents being exemplary.

R^(d)′, R^(d)″ and R^(d)′″ each independently refer to hydrogen,unsubstituted (C₁-C₈)alkyl, unsubstituted hetero(C₁-C₈) alkyl,unsubstituted aryl and aryl substituted with one to three substituentsselected from -halo, unsubstituted alkyl, unsubstituted alkoxy,unsubstituted thioalkoxy and unsubstituted aryl (C₁-C₄)alkyl. WhenR^(d)′ and R^(d)″ are attached to the same nitrogen atom, they can becombined with the nitrogen atom to form a 5-, 6- or 7-membered ring. Forexample, —NR^(d)′R^(d)″ can represent 1-pyrrolidinyl or 4-morpholinyl.

Typically, an alkyl or heteroalkyl group will have from zero to threesubstituents, with those groups having two or fewer substituents beingexemplary of the present invention. An alkyl or heteroalkyl radical canbe unsubstituted or monosubstituted. In some embodiments, an alkyl orheteroalkyl radical will be unsubstituted.

Exemplary substituents for the alkyl and heteroalkyl radicals includebut are not limited to —OR^(d)′, ═O, ═NR^(d)′, ═N—OR^(d)′,—NR^(d)′R^(d)″, —SR^(d)′, -halo, —SiR^(d)′R^(d)″R^(d)′″, —OC(O)R^(d)′,—C(O)R^(d)′, —CO₂R^(d)′, —CONR^(d)′R^(d)″, —OC(O)NR^(d)′R^(d)″,—NR^(d)″C(O)R^(d)′, —NR^(d)′″C(O)NR^(d)′R^(d)″,—NR^(d)′″SO₂NR^(d)′R^(d)″, —NR^(d)″CO₂R^(d)′, —NHC(NH₂)═NH,—NR^(a)′C(NH₂)═NH, —NHC(NH₂)═NR^(d)′, —S(O)R^(d)′, —SO₂R^(d)′,—SO₂NR^(d)′ R^(d)″, —NR^(d)″SO₂R^(d)′, —CN and —NO₂, where R^(d)′,R^(d)″ and R^(d)′″ are as defined above. Typical substituents can beselected from: —OR^(d), ═O, —NR^(d)′R^(d)″, -halo, —OC(O)R^(d)′,—CO₂R^(d)′, —C(O)NR^(d)′R^(d)″, —OC(O)NR^(d)′R^(d)″, —NR^(d)″C(O)R^(d)′,—NR^(d)″CO₂R^(d)′, —NR^(d)′″SO₂NR^(d)′R^(d)″, —SO₂R^(d)′,—SO₂NR^(d)′R^(d)″, —NR^(d)″SO₂R^(d)′—CN and —NO₂.

Similarly, substituents for the aryl and heteroaryl groups are variedand selected from: -halo, —OR^(e)′, —OC(O)R^(e)′, —NR^(e)′R^(e)″,—SR^(e)′, —R^(e)′, —CN, —NO₂, —CO₂R^(e)′, —NR^(e)′″C(O)NR^(e)′R^(e)″,—C(O)R^(e)′, —OC(O)NR^(e)′R^(e)″, —NR^(e)″C(O)R^(e)′, —NR^(e)″CO₂R^(e)′,—NR^(e)′″C(O)NR^(e)′R^(e)″, —NR^(e)′″SO₂NR^(e)′R^(e)″, —NHC(NH₂)═NH,—NR^(e)′C(NH₂)═NH, —NH—C(NH₂)═NR^(e)′, —S(O)R^(e)′, —SO₂R^(e)′,—SO₂NR^(e)′R^(e)″, —NR^(e)″SO₂R^(e)′, —N₃, —CH(Ph)₂, perfluoroalkoxy andperfluoro(C₁-C₄)alkyl, in a number ranging from zero to the total numberof open valences on the aromatic ring system.

R^(e)′, R^(e)″ and R^(e)′″ are independently selected from hydrogen,unsubstituted (C₁-C₈) alkyl, unsubstituted hetero(C₁-C₈) alkyl,unsubstituted aryl, unsubstituted heteroaryl, unsubstituted aryl(C₁-C₄)alkyl and unsubstituted aryloxy(C₁-C₄) alkyl. Typically, an aryl orheteroaryl group will have from zero to three substituents, with thosegroups having two or fewer substituents being exemplary in the presentinvention. In one embodiment of the invention, an aryl or heteroarylgroup will be unsubstituted or monosubstituted. In another embodiment,an aryl or heteroaryl group will be unsubstituted.

Two of the substituents on adjacent atoms of an aryl or heteroaryl ringin an aryl or heteroaryl group as described herein may optionally bereplaced with a substituent of the formula -T-C(O)—(CH₂)_(q)—U—, whereinT and U are independently —NH—, —O—, —CH₂— or a single bond, and q is aninteger of from 0 to 2. Alternatively, two of the substituents onadjacent atoms of the aryl or heteroaryl ring may optionally be replacedwith a substituent of the formula -J-(CH₂)_(r)—K—, wherein J and K areindependently —CH₂—, —O—, —NH—, —S—, —S(O)—, —S(O)₂—, —S(O)₂NR^(f)′- ora single bond, and r is an integer of from 1 to 3. One of the singlebonds of the new ring so formed may optionally be replaced with a doublebond. Alternatively, two of the substituents on adjacent atoms of thearyl or heteroaryl ring may optionally be replaced with a substituent ofthe formula —(CH₂)_(s)—X—(CH₂)_(t)—, where s and t are independentlyintegers of from 0 to 3, and X is —O—, —NR^(f)′—, —S—, —S(O)—, —S(O)₂—,or —S(O)₂NR^(a)′—. The substituent R^(f)′ in —NR^(f)′— and—S(O)₂NR^(f)′— is selected from hydrogen or unsubstituted (C₁-C₆) alkyl.

“Stable compound” and “stable structure” are meant to indicate acompound that is sufficiently robust to survive isolation to a usefuldegree of purity from a reaction mixture, and formulation into anefficacious therapeutic agent.

As used herein the term “therapeutically effective amount” generallymeans the amount necessary to ameliorate at least one symptom of adisorder to be prevented, reduced, or treated as described herein. Thephrase “therapeutically effective amount” as it relates to the GSNORinhibitors of the present invention shall mean the GSNOR inhibitordosage that provides the specific pharmacological response for which theGSNOR inhibitor is administered in a significant number of subjects inneed of such treatment. It is emphasized that a therapeuticallyeffective amount of a GSNOR inhibitor that is administered to aparticular subject in a particular instance will not always be effectivein treating the conditions/diseases described herein, even though suchdosage is deemed to be a therapeutically effective amount by those ofskill in the art. The phrase “therapeutically effective amount” as itrelates to NK3 receptor antagonists of the present invention shall meanthe NK3 receptor antagonist dosage that provides the specificpharmacological response for which the NK3 receptor antagonist isadministered in a significant number of subjects in need of suchtreatment. It is emphasized that a therapeutically effective amount of aNK3 receptor antagonist that is administered to a particular subject ina particular instance will not always be effective in treating theconditions/diseases described herein, even though such dosage is deemedto be a therapeutically effective amount by those of skill in the art.

The term “biological sample” includes, but is not limited to, samples ofblood (e.g., serum, plasma, or whole blood), urine, saliva, sweat,breast milk, vaginal secretions, semen, hair follicles, skin, teeth,bones, nails, or other secretions, body fluids, tissues, or cells. Inaccordance with the invention, the levels of the S-nitrosoglutathionereductase in the biological sample can be determined by the methodsdescribed in U.S. Patent Application Publication No. 2005/0014697.

D. Pharmaceutical Compositions

The invention encompasses pharmaceutical compositions comprising atleast one compound of the invention described herein and at least onepharmaceutically acceptable carrier. Suitable carriers are described in“Remington: The Science and Practice, Twentieth Edition,” published byLippincott Williams & Wilkins, which is incorporated herein byreference. Pharmaceutical compositions according to the invention mayalso comprise one or more non-inventive compound active agents.

The pharmaceutical compositions of the invention can comprise novelcompounds described herein, the pharmaceutical compositions can compriseknown compounds which previously were not know to have GSNOR inhibitoractivity or NK3 receptor antagonist activity, or a combination thereof.

The compounds of the invention can be utilized in any pharmaceuticallyacceptable dosage form, including but not limited to injectable dosageforms, liquid dispersions, gels, aerosols, ointments, creams,lyophilized formulations, dry powders, tablets, capsules, controlledrelease formulations, fast melt formulations, delayed releaseformulations, extended release formulations, pulsatile releaseformulations, mixed immediate release and controlled releaseformulations, etc. Specifically, the compounds of the inventiondescribed herein can be formulated: (a) for administration selected fromthe group consisting of oral, pulmonary, intravenous, intra-arterial,intrathecal, intra-articular, rectal, ophthalmic, colonic, parenteral,intracisternal, intravaginal, intraperitoneal, local, buccal, nasal, andtopical administration; (b) into a dosage form selected from the groupconsisting of liquid dispersions, gels, aerosols, ointments, creams,tablets, sachets and capsules; (c) into a dosage form selected from thegroup consisting of lyophilized formulations, dry powders, fast meltformulations, controlled release formulations, delayed releaseformulations, extended release formulations, pulsatile releaseformulations, and mixed immediate release and controlled releaseformulations; or (d) any combination thereof.

For respiratory infections, an inhalation formulation can be used toachieve high local concentrations. Formulations suitable for inhalationinclude dry power or aerosolized or vaporized solutions, dispersions, orsuspensions capable of being dispensed by an inhaler or nebulizer intothe endobronchial or nasal cavity of infected patients to treat upperand lower respiratory bacterial infections.

Solutions or suspensions used for parenteral, intradermal, orsubcutaneous application can comprise one or more of the followingcomponents: (1) a sterile diluent such as water for injection, salinesolution, fixed oils, polyethylene glycols, glycerine, propylene glycolor other synthetic solvents; (2) antibacterial agents such as benzylalcohol or methyl parabens; (3) antioxidants such as ascorbic acid orsodium bisulfite; (4) chelating agents such asethylenediaminetetraacetic acid; (5) buffers such as acetates, citratesor phosphates; and (5) agents for the adjustment of tonicity such assodium chloride or dextrose. The pH can be adjusted with acids or bases,such as hydrochloric acid or sodium hydroxide. A parenteral preparationcan be enclosed in ampoules, disposable syringes or multiple dose vialsmade of glass or plastic.

Pharmaceutical compositions suitable for injectable use may comprisesterile aqueous solutions (where water soluble) or dispersions andsterile powders for the extemporaneous preparation of sterile injectablesolutions or dispersion. For intravenous administration, suitablecarriers include physiological saline, bacteriostatic water, CremophorEL (BASF, Parsippany, N.J.) or phosphate buffered saline (PBS). In allcases, the composition must be sterile and should be fluid to the extentthat easy syringability exists. The pharmaceutical composition should bestable under the conditions of manufacture and storage and should bepreserved against the contaminating action of microorganisms such asbacteria and fungi.

The carrier can be a solvent or dispersion medium comprising, forexample, water, ethanol, polyol (for example, glycerol, propyleneglycol, and liquid polyethylene glycol, and the like), and suitablemixtures thereof. The proper fluidity can be maintained, for example, bythe use of a coating such as lecithin, by the maintenance of therequired particle size in the case of dispersion and by the use ofsurfactants. Prevention of the action of microorganisms can be achievedby various antibacterial and antifungal agents, for example, parabens,chlorobutanol, phenol, ascorbic acid, thimerosal, and the like. In manycases, it will be preferable to include isotonic agents, for example,sugars, polyalcohols such as manitol or sorbitol, and inorganic saltssuch as sodium chloride in the composition. Prolonged absorption of theinjectable compositions can be brought about by including in thecomposition an agent which delays absorption, for example, aluminummonostearate and gelatin.

Sterile injectable solutions can be prepared by incorporating the activereagent in the required amount in an appropriate solvent with one or acombination of ingredients enumerated above, as required, followed byfiltered sterilization. Generally, dispersions are prepared byincorporating at least one compound of the invention into a sterilevehicle that contains a basic dispersion medium and any other requiredingredients. In the case of sterile powders for the preparation ofsterile injectable solutions, exemplary methods of preparation includevacuum drying and freeze-drying, both of which yield a powder of acompound of the invention plus any additional desired ingredient from apreviously sterile-filtered solution thereof.

Oral compositions generally include an inert diluent or an ediblecarrier. They can be enclosed, for example, in gelatin capsules orcompressed into tablets. For the purpose of oral therapeuticadministration, the compound of the invention can be incorporated withexcipients and used in the form of tablets, troches, or capsules. Oralcompositions can also be prepared using a fluid carrier for use as amouthwash, wherein the compound in the fluid carrier is applied orallyand swished and expectorated or swallowed. Pharmaceutically compatiblebinding agents, and/or adjuvant materials can be included as part of thecomposition.

For administration by inhalation, the compounds are delivered in theform of an aerosol spray from pressured container or dispenser thatcontains a suitable propellant, e.g., a gas such as carbon dioxide, anebulized liquid, or a dry powder from a suitable device. Fortransmucosal or transdermal administration, penetrants appropriate tothe barrier to be permeated are used in the formulation. Such penetrantsare generally known in the art, and include, for example, fortransmucosal administration, detergents, bile salts, and fusidic acidderivatives. Transmucosal administration can be accomplished through theuse of nasal sprays or suppositories. For transdermal administration,the active reagents are formulated into ointments, salves, gels, orcreams as generally known in the art. The reagents can also be preparedin the form of suppositories (e.g., with conventional suppository basessuch as cocoa butter and other glycerides) or retention enemas forrectal delivery.

In one embodiment, the compounds of the invention are prepared withcarriers that will protect against rapid elimination from the body. Forexample, a controlled release formulation can be used, includingimplants and microencapsulated delivery systems. Biodegradable,biocompatible polymers can be used, such as ethylene vinyl acetate,polyanhydrides, polyglycolic acid, collagen, polyorthoesters, andpolylactic acid. Methods for preparation of such formulations will beapparent to those skilled in the art.

Liposomal suspensions (including liposomes targeted to infected cellswith monoclonal antibodies to viral antigens) can also be used aspharmaceutically acceptable carriers. These can be prepared according tomethods known to those skilled in the art, for example, as described inU.S. Pat. No. 4,522,811.

Additionally, suspensions of the compounds of the invention may beprepared as appropriate oily injection suspensions. Suitable lipophilicsolvents or vehicles include fatty oils, such as sesame oil, orsynthetic fatty acid esters, such as ethyl oleate, triglycerides, orliposomes. Non-lipid polycationic amino polymers may also be used fordelivery. Optionally, the suspension may also include suitablestabilizers or agents to increase the solubility of the compounds andallow for the preparation of highly concentrated solutions.

It is especially advantageous to formulate oral or parenteralcompositions in dosage unit form for ease of administration anduniformity of dosage. Dosage unit form as used herein refers tophysically discrete units suited as unitary dosages for the subject tobe treated; each unit containing a predetermined quantity of thecompound of the invention calculated to produce the desired therapeuticeffect in association with the required pharmaceutical carrier. Thespecification for the dosage unit forms of the invention are dictated byand directly dependent on the unique characteristics of the compound ofthe invention and the particular therapeutic effect to be achieved, andthe limitations inherent in the art of compounding such an active agentfor the treatment of individuals.

Pharmaceutical compositions according to the invention comprising atleast one compound of the invention can comprise one or morepharmaceutical excipients. Examples of such excipients include, but arenot limited to binding agents, filling agents, lubricating agents,suspending agents, sweeteners, flavoring agents, preservatives, buffers,wetting agents, disintegrants, effervescent agents, and otherexcipients. Such excipients are known in the art. Exemplary excipientsinclude: (1) binding agents which include various celluloses andcross-linked polyvinylpyrrolidone, microcrystalline cellulose, such asAvicel® PH101 and Avicel® PH102, silicified microcrystalline cellulose(ProSolv SMCC™), gum tragacanth and gelatin; (2) filling agents such asvarious starches, lactose, lactose monohydrate, and lactose anhydrous;(3) disintegrating agents such as alginic acid, Primogel, corn starch,lightly crosslinked polyvinyl pyrrolidone, potato starch, maize starch,and modified starches, croscarmellose sodium, cross-povidone, sodiumstarch glycolate, and mixtures thereof; (4) lubricants, including agentsthat act on the flowability of a powder to be compressed, includemagnesium stearate, colloidal silicon dioxide, such as Aerosil® 200,talc, stearic acid, calcium stearate, and silica gel; (5) glidants suchas colloidal silicon dioxide; (6) preservatives, such as potassiumsorbate, methylparaben, propylparaben, benzoic acid and its salts, otheresters of parahydroxybenzoic acid such as butylparaben, alcohols such asethyl or benzyl alcohol, phenolic compounds such as phenol, orquaternary compounds such as benzalkonium chloride; (7) diluents such aspharmaceutically acceptable inert fillers, such as microcrystallinecellulose, lactose, dibasic calcium phosphate, saccharides, and/ormixtures of any of the foregoing; examples of diluents includemicrocrystalline cellulose, such as Avicel® PH101 and Avicel® PH102;lactose such as lactose monohydrate, lactose anhydrous, and Pharmatose®DCL21; dibasic calcium phosphate such as Emcompress; mannitol; starch;sorbitol; sucrose; and glucose; (8) sweetening agents, including anynatural or artificial sweetener, such as sucrose, saccharin sucrose,xylitol, sodium saccharin, cyclamate, aspartame, and acesulfame; (9)flavoring agents, such as peppermint, methyl salicylate, orangeflavoring, Magnasweet® (trademark of MAFCO), bubble gum flavor, fruitflavors, and the like; and (10) effervescent agents, includingeffervescent couples such as an organic acid and a carbonate orbicarbonate. Suitable organic acids include, for example, citric,tartaric, malic, fumaric, adipic, succinic, and alginic acids andanhydrides and acid salts. Suitable carbonates and bicarbonates include,for example, sodium carbonate, sodium bicarbonate, potassium carbonate,potassium bicarbonate, magnesium carbonate, sodium glycine carbonate,L-lysine carbonate, and arginine carbonate. Alternatively, only thesodium bicarbonate component of the effervescent couple may be present.

E. Kits Comprising the Compositions of the Invention

The present invention also encompasses kits comprising the compositionsof the invention. Such kits can comprise, for example, (1) at least onecompound of the invention; and (2) at least one pharmaceuticallyacceptable carrier, such as a solvent or solution. Additional kitcomponents can optionally include, for example: (1) any of thepharmaceutically acceptable excipients identified herein, such asstabilizers, buffers, etc., (2) at least one container, vial or similarapparatus for holding and/or mixing the kit components; and (3) deliveryapparatus, such as an inhaler, nebulizer, syringe, etc.

F. Methods of Preparing Compounds of the Invention

The compounds of the invention can readily be synthesized using knownsynthetic methodologies or via a modification of known syntheticmethodologies. As would be readily recognized by a skilled artisan, themethodologies described below allow the synthesis ofdihydropyridin-2(1H)-ones having a variety of substituents. Exemplarysynthetic methods are described in the examples below.

If needed, further purification and separation of enantiomers anddiastereomers can be achieved by routine procedures known in the art.Thus, for example, the separation of enantiomers of a compound can beachieved by the use of chiral HPLC and related chromatographictechniques. Diastereomers can be similarly separated. In some instances,however, diastereomers can simply be separated physically, such as, forexample, by controlled precipitation or crystallization.

The process of the invention, when carried out as prescribed herein, canbe conveniently performed at temperatures that are routinely accessiblein the art. In one embodiment, the process is performed at a temperaturein the range of about 25° C. to about 110° C. In another embodiment, thetemperature is in the range of about 40° C. to about 100° C. In yetanother embodiment, the temperature is in the range of about 50° C. toabout 95° C.

Synthetic steps that require a base are carried out using any convenientorganic or inorganic base. Typically, the base is not nucleophilic.Thus, in one embodiment, the base is selected from carbonates,phosphates, hydroxides, alkoxides, salts of disilazanes, and tertiaryamines.

The process of the invention, when performed as described herein, can besubstantially complete after several minutes to after several hoursdepending upon the nature and quantity of reactants and reactiontemperature. The determination of when the reaction is substantiallycomplete can be conveniently evaluated by ordinary techniques known inthe art such as, for example, HPLC, LCMS, TLC, and ¹H NMR.

G. Methods of Treatment

The invention encompasses methods of preventing or treating (e.g.,alleviating one or more symptoms of) medical conditions through use ofone or more of the disclosed compounds. The methods compriseadministering a therapeutically effective amount of a compound of theinvention to a patient in need. The compositions of the invention canalso be used for prophylactic therapy.

The compound of the invention used in the methods of treatment accordingto the invention can be: (1) a novel compound described herein, or apharmaceutically acceptable salt thereof, a prodrug thereof, or ametabolite thereof; (2) a compound which was known prior to the presentinvention, but wherein it was not known that the compound is a GSNORinhibitor or NK3 receptor antagonist, or a pharmaceutically acceptablesalt thereof, a prodrug thereof, or a metabolite thereof; or (3) acompound which was known prior to the present invention, and wherein itwas known that the compound is a GSNOR inhibitor or NK3 receptorantagonist, but wherein it was not known that the compound is useful forthe methods of treatment described herein, or a pharmaceuticallyacceptable salt thereof, a prodrug thereof, or a metabolite thereof.

The patient can be any animal, domestic, livestock or wild, including,but not limited to cats, dogs, horses, pigs and cattle, and preferablyhuman patients. As used herein, the terms patient and subject may beused interchangeably.

As used herein, “treating” describes the management and care of apatient for the purpose of combating a disease, condition, or disorderand includes the administration of a compound of the present inventionto prevent the onset of the symptoms or complications, alleviating thesymptoms or complications, or eliminating the disease, condition ordisorder. More specifically, “treating” includes reversing, attenuating,alleviating, minimizing, suppressing or halting at least one deleterioussymptom or effect of a disease (disorder) state, disease progression,disease causative agent (e.g., bacteria or viruses), or other abnormalcondition. Treatment is continued as long as symptoms and/or pathologyameliorate.

In general, the dosage, i.e., the therapeutically effective amount,ranges from 1 μg to 10 g/kg and often ranges from 10 μg to 1 g/kg or 10μg to 100 mg/kg body weight of the subject being treated, per day.

H. GSNOR Uses

In subjects with deleteriously high levels of GSNOR or GSNOR activity,modulation may be achieved, for example, by administering one or more ofthe disclosed compounds that disrupts or down-regulates GSNOR function,or decreases GSNOR levels. These compounds may be administered withother GSNOR inhibitor agents, such as anti-GSNOR antibodies or antibodyfragments, GSNOR antisense, iRNA, or small molecules, or otherinhibitors, alone or in combination with other agents as described indetail herein.

The present invention provides a method of treating a subject afflictedwith a disorder ameliorated by NO donor therapy. Such a method comprisesadministering to a subject a therapeutically effective amount of a GSNORinhibitor.

The disorders can include pulmonary disorders associated with hypoxemiaand/or smooth muscle constriction in the lungs and/or lung infectionand/or lung injury (e.g., pulmonary hypertension, ARDS, asthma,pneumonia, pulmonary fibrosis/interstitial lung diseases, cysticfibrosis, COPD) cardiovascular disease and heart disease, includingconditions such as hypertension, ischemic coronary syndromes,atherosclerosis, heart failure, glaucoma, diseases characterized byangiogenesis (e.g., coronary artery disease), disorders where there isrisk of thrombosis occurring, disorders where there is risk ofrestenosis occurring, chronic inflammatory diseases (e.g., AID dementiaand psoriasis), diseases where there is risk of apoptosis occurring(e.g., heart failure, atherosclerosis, degenerative neurologicdisorders, arthritis and liver injury (e.g., ischemic or alcoholic)),impotence, obesity caused by eating in response to craving for foodstroke, reperfusion injury (e.g., traumatic muscle injury in heart orlung or crush injury), and disorders where preconditioning of heart orbrain for NO protection against subsequent ischemic events isbeneficial.

In one embodiment, the compounds of the present invention or apharmaceutically acceptable salt thereof, or a prodrug or metabolitethereof, can be administered in combination with an NO donor. An NOdonor donates nitric oxide or a related redox species and more generallyprovides nitric oxide bioactivity, that is activity which is identifiedwith nitric oxide, e.g., vasorelaxation or stimulation or inhibition ofa receptor protein, e.g., ras protein, adrenergic receptor, NFκB. NOdonors including S-nitroso, O-nitroso, C-nitroso and N-nitroso compoundsand nitro derivatives thereof and metal NO complexes, but not excludingother NO bioactivity generating compounds, useful herein are describedin “Methods in Nitric Oxide Research,” Feelisch et al. eds., pages71-115 (J. S., John Wiley & Sons, New York, 1996), which is incorporatedherein by reference. NO donors which are C-nitroso compounds wherenitroso is attached to a tertiary carbon which are useful herein includethose described in U.S. Pat. No. 6,359,182 and in WO 02/34705. Examplesof S-nitroso compounds, including S-nitrosothiols useful herein,include, for example, S-nitrosoglutathione,S-nitroso-N-acetylpenicillamine, S-nitroso-cysteine and ethyl esterthereof, S-nitroso cysteinyl glycine,S-nitroso-gamma-methyl-L-homocysteine, S-nitroso-L-homocysteine,S-nitroso-gamma-thio-L-leucine, S-nitroso-delta-thio-L-leucine, andS-nitrosoalbumin. Examples of other NO donors useful herein are sodiumnitroprusside (nipride), ethyl nitrite, isosorbide, nitroglycerin, SIN 1which is molsidomine, furoxamines, N-hydroxy (N-nitrosamine) andperfluorocarbons that have been saturated with NO or a hydrophobic NOdonor.

The combination of a GSNOR inhibitor with R(+) enantiomer of amlodipine,a known NO releaser (Zhang X. P at al. 2002 J. CardiovascularPharmacology 39, 208-214) is also an embodiment of the presentinvention.

The present invention also provides a method of treating a subjectafflicted with pathologically proliferating cells where the methodcomprises administering to said subject a therapeutically effectiveamount of an inhibitor of GSNOR. The inhibitors of GSNOR are thecompounds as defined above, or a pharmaceutically acceptable saltthereof, or a prodrug or metabolite thereof, in combination with apharmaceutically acceptable carrier. Treatment is continued as long assymptoms and/or pathology ameliorate.

In another embodiment, the pathologically proliferating cells can bepathologically proliferating microbes. The microbes involved can bethose where GSNOR is expressed to protect the microbe from nitrosativestress or where a host cell infected with the microbe expresses theenzyme, thereby protecting the microbe from nitrosative stress. The term“pathologically proliferating microbes” is used herein to meanpathologic microorganisms including but not limited to pathologicbacteria, pathologic viruses, pathologic Chlamydia, pathologic protozoa,pathologic Rickettsia, pathologic fungi, and pathologic mycoplasmata.More detail on the applicable microbes is set forth at columns 11 and 12of U.S. Pat. No. 6,057,367. The term “host cells infected withpathologic microbes” includes not only mammalian cells infected withpathologic viruses but also mammalian cells containing intracellularbacteria or protozoa, e.g., macrophages containing Mycobacteriumtuberculosis, Mycobacterium leper (leprosy), or Salmonella typhi(typhoid fever).

In another embodiment, the pathologically proliferating cells can bepathologic helminths. The term “pathologic helminths” is used herein torefer to pathologic nematodes, pathologic trematodes and pathologiccestodes. More detail on the applicable helminths is set forth at column12 of U.S. Pat. No. 6,057,367.

In another embodiment, the pathologically proliferating cells can bepathologically proliferating mammalian cells. The term “pathologicallyproliferating mammalian cells” as used herein means cells of the mammalthat grow in size or number in said mammal so as to cause a deleteriouseffect in the mammal or its organs. The term includes, for example, thepathologically proliferating or enlarging cells causing restenosis, thepathologically proliferating or enlarging cells causing benign prostatichypertrophy, the pathologically proliferating cells causing myocardialhypertrophy and proliferating cells at inflammatory sites such assynovial cells in arthritis or cells associated with a cellproliferation disorder.

As used herein, the term “cell proliferative disorder” refers toconditions in which the unregulated and/or abnormal growth of cells canlead to the development of an unwanted condition or disease, which canbe cancerous or non-cancerous, for example a psoriatic condition. Asused herein, the term “psoriatic condition” refers to disordersinvolving keratinocyte hyperproliferation, inflammatory cellinfiltration, and cytokine alteration. The cell proliferative disordercan be a precancerous condition or cancer. The cancer can be primarycancer or metastatic cancer, or both.

As used herein, the term “cancer” includes solid tumors, such as lung,breast, colon, ovarian, pancreas, prostate, adenocarcinoma, squamouscarcinoma, sarcoma, malignant glioma, leiomyosarcoma, hepatoma, head andneck cancer, malignant melanoma, non-melanoma skin cancers, as well ashematologic tumors and/or malignancies, such as leukemia, childhoodleukemia and lymphomas, multiple myeloma, Hodgkin's disease, lymphomasof lymphocytic and cutaneous origin, acute and chronic leukemia such asacute lymphoblastic, acute myelocytic or chronic myelocytic leukemia,plasma cell neoplasm, lymphoid neoplasm and cancers associated withAIDS.

In addition to psoriatic conditions, the types of proliferative diseaseswhich may be treated using the compositions of the present invention areepidermic and dermoid cysts, lipomas, adenomas, capillary and cutaneoushemangiomas, lymphangiomas, nevi lesions, teratomas, nephromas,myofibromatosis, osteoplastic tumors, and other dysplastic masses andthe like. In one embodiment, proliferative diseases include dysplasiasand disorders of the like.

In one embodiment, the treating cancer comprises a reduction in tumorsize, decrease in tumor number, a delay of tumor growth, decrease inmetastaic lesions in other tissues or organs distant from the primarytumor site, an improvement in the survival of patients, or animprovement in the quality of patient life, or at least two of theabove.

In another embodiment, the treating a cell proliferative disordercomprises a reduction in the rate of cellular proliferation, reductionin the proportion of proliferating cells, a decrease in size of an areaor zone of cellular proliferation, or a decrease in the number orproportion of cells having an abnormal appearance or morphology, or atleast two of the above.

In yet another embodiment, the compounds of the present invention or apharmaceutically acceptable salt thereof, a prodrug thereof, ormetabolite thereof, can be administered in combination with a secondchemotherapeutic agent. In a further embodiment, the secondchemotherapeutic agent is selected from the group consisting oftamoxifen, raloxifene, anastrozole, exemestane, letrozole, cisplatin,carboplatin, paclitaxel, cyclophosphamide, lovastatin, minosine,gemcitabine, araC, 5-fluorouracil, methotrexate, docetaxel, goserelin,vincristin, vinblastin, nocodazole, teniposide, etoposide, epothilone,navelbine, camptothecin, daunonibicin, dactinomycin, mitoxantrone,amsacrine, doxorubicin, epirubicin, idarubicin imatanib, gefitinib,erlotinib, sorafenib, sunitinib malate, trastuzumab, rituximab,cetuximab, and bevacizumab.

In one embodiment, the compounds of the present invention or apharmaceutically acceptable salt thereof, a prodrug thereof, ormetabolite thereof, can be administered in combination with an agentthat imposes nitrosative or oxidative stress. Agents for selectivelyimposing nitrosative stress to inhibit proliferation of pathologicallyproliferating cells in combination therapy with GSNOR inhibitors hereinand dosages and routes of administration therefor include thosedisclosed in U.S. Pat. No. 6,057,367, which is incorporated herein.Supplemental agents for imposing oxidative stress (i.e., agents thatincrease GSSG (oxidized glutathione) over GSH (glutathione) ratio orNAD(P) over NAD(P)H ratio or increase thiobarbituric acid derivatives)in combination therapy with GS-FDH inhibitors herein include, forexample, L-buthionine-S-sulfoximine (BSO), glutathione reductaseinhibitors (e.g., BCNU), inhibitors or uncouplers of mitochondrialrespiration and drugs that increase reactive oxygen species (ROS), e.g.,adriamycin, in standard dosages with standard routes of administration.

GSNOR inhibitors may also be co-administered with a phosphodiesteraseinhibitor (e.g., rolipram, cilomilast, roflumilast, Viagra® (sildenifilcitrate), Cialis® (tadalafil), Levitra® (vardenifil), etc.), aβ-agonist, a steroid, or a leukotriene antagonist (LTD-4). Those skilledin the art can readily determine the appropriate therapeuticallyeffective amount depending on the disorder to be ameliorated.

GSNOR inhibitors may be used as a means to improve β-adrenergicsignaling. In particular, inhibitors of GSNOR alone or in combinationwith β-agonists could be used to treat or protect against heart failure,or other vascular disorders such as hypertension and asthma. GSNORinhibitors can also be used to modulate G protein coupled receptors(GPCRs) by potentiating Gs G-protein, leading to smooth musclerelaxation (e.g., airway and blood vessels), and by attenuating GqG-protein, and thereby preventing smooth muscle contraction (e.g., inairway and blood vessels).

The therapeutically effective amount for the treatment of a subjectafflicted with a disorder ameliorated by NO donor therapy is the GSNORinhibiting amount in vivo that causes amelioration of the disorder beingtreated or protects against a risk associated with the disorder. Forexample, for asthma, a therapeutically effective amount is abronchodilating effective amount; for cystic fibrosis, a therapeuticallyeffective amount is an airway obstruction ameliorating effective amount;for ARDS, a therapeutically effective amount is a hypoxemia amelioratingeffective amount; for heart disease, a therapeutically effective amountis an angina relieving or angiogenesis inducing effective amount; forhypertension, a therapeutically effective amount is a blood pressurereducing effective amount; for ischemic coronary disorders, atherapeutic amount is a blood flow increasing effective amount; foratherosclerosis, a therapeutically effective amount is an endothelialdysfunction reversing effective amount; for glaucoma, a therapeuticamount is an intraocular pressure reducing effective amount; fordiseases characterized by angiogenesis, a therapeutically effectiveamount is an angiogenesis inhibiting effective amount; for disorderswhere there is risk of thrombosis occurring, a therapeutically effectiveamount is a thrombosis preventing effective amount; for disorders wherethere is risk of restenosis occurring, a therapeutically effectiveamount is a restenosis inhibiting effective amount; for chronicinflammatory diseases, a therapeutically effective amount is aninflammation reducing effective amount; for disorders where there isrisk of apoptosis occurring, a therapeutically effective amount is anapoptosis preventing effective amount; for impotence, a therapeuticallyeffective is an erection attaining or sustaining effective amount; forobesity, a therapeutically effective amount is a satiety causingeffective amount; for stroke, a therapeutically effective amount is ablood flow increasing or a TIA protecting effective amount; forreperfusion injury, a therapeutically effective amount is a functionincreasing effective amount; and for preconditioning of heart and brain,a therapeutically effective amount is a cell protective effectiveamount, e.g., as measured by triponin or CPK.

The therapeutically effective amount for the treatment of a subjectafflicted with pathologically proliferating cells means a GSNORinhibiting amount in vivo which is an antiproliferative effectiveamount. Such antiproliferative effective amount as used herein means anamount causing reduction in rate of proliferation of at least about 20%,at least about 10%, at least about 5%, or at least about 1%.

I. NK3 Uses

The compounds of Formula I, particularly the active enantiomers thatantagonize the NK3 receptor are useful in the prevention and treatmentof a wide variety of clinical conditions which are characterized byoverstimulation of the tachkinin receptors, in particular, NK1, NK2 andNK3, and most particularly NK3. These conditions may include disordersof the central nervous system (CNS) such as anxiety, depression,psychosis, and schizophrenia; neurodegenerative disorders such as AIDSrelated dementia, senile dementia of the Alzheimer's type, Alzheimer'sdisease and Down's syndrome; demyelinating diseases such as multiplesclerosis and amyotrophic lateral sclerosis and other neuropathologicaldisorders such as diabetic or peripheral neuropathy, AIDS relatedneuropathy, chemotherapy-induced neuropathy, and neuralgia; respiratorydiseases such as chronic obstructive airways disease, bronchopneumonia,bronchospasm and asthma; inflammatory diseases such as inflammatorybowel disease, psoriasis, fibrositis, osteoarthritis, and rheumatoidarthritis; allergies such as eczema and rhinitis; hypersensitivitydisorders such as poison ivy; ophthalmic diseases such asconjunctivitis, vernal conjunctivitis, and the like; cutaneous diseasessuch as contact dermatitis, atopic dermatitis, urticaria, and othereczemoatoid dermatitis; addiction disorders such as alcoholism; stressrelated somatic disorders; reflex sympathetic dystrophy such asshoulder/had syndrome; dysthymic disorders; adverse immunologicalreactions such as rejection of transplanted tissues and disordersrelated to immune enhancement or suppression such as systemic lupuserythematosis; gastrointestinal (GI) disorders and diseases of the GItract such as disorders associated with the neuronal control of viscerasuch as ulcerative colitis, Crohn's disease and incontinence; disordersof bladder function; fibrosing and collagen diseases such as sclerodermaand eosinophilic fascioliasis; disorders of blood flow caused byvasodilation and vasospastic diseases such as angina, migraine andReynaud's disease; and pain or nociception, for example, that isattributable to or associated with any of the foregoing conditionsespecially the transmission of pain in migraine. Hence, these compoundsare readily adapted to therapeutic use for the treatment ofphysiological disorders associated with the overstimulation of thetachykinin receptors, in particular NK1, NK2 and NK3, and mostparticularly NK3.

J. Uses in an Apparatus

The compounds of the present invention or a pharmaceutically acceptablesalt thereof, or a prodrug or metabolite thereof, can be applied tovarious apparatus in circumstances when the presence of such compoundswould be beneficial. Such apparatus can be any device or container, forexample, implantable devices in which a compound of the invention can beused to coat a surgical mesh or cardiovascular stent prior toimplantation in a patient. The compounds of the invention can also beapplied to various apparatus for in vitro assay purposes or forculturing cells.

The compounds of the present invention or a pharmaceutically acceptablesalt thereof, or a prodrug or metabolite thereof, can also be used as anagent for the development, isolation or purification of binding partnersto compounds of the invention, such as antibodies, natural ligands, andthe like. Those skilled in the art can readily determine related usesfor the compounds of the present invention.

EXAMPLES

The following examples are given to illustrate the present invention. Itshould be understood, however, that the invention is not to be limitedto the specific conditions or details described in these examples.Throughout the specification, any and all references to a publiclyavailable document, including a U.S. patent, are specificallyincorporated by reference.

Examples 1-17 list representative novel dihydropyridin-2(1H)-one analogsof Formula I useful as GSNOR inhibitors and NK3 antagonists of theinvention. The synthetic methods that can be used to prepare eachcompound are detailed in Examples 1-17. In some cases, the startingmaterial was not commercially available. In these cases, the synthesisof the intermediates is described in Example 18. Supporting massspectrometry data and proton NMR data for each compound is also includedin Examples 1-17.

Example 1 Compound 1,4-(3-ethoxy-4-hydroxy-5-nitrophenyl)-5,6-diphenyl-3,4-dihydropyridin-2(1H)-one

A mixture of 1,2-diphenylethanone (150 mg, 0.76 mmol),3-ethoxy-4-hydroxy-5-nitrobenzaldehyde (194 mg, 0.92 mmol), meldrum'sacid (132 mg, 0.92 mmol) and ammonium acetate (71 mg, 0.92 mmol) inacetic acid (5 mL) was refluxed overnight. The solvent was removed underreduced pressure and the residue was purified by silica gel columnchromatography (EtOAc: PE=2:1) and prep-HPLC (0.1% TFA as additive) toafford Compound 1 as yellow solid (40 mg, yield: 13%). ¹H NMR (DMSO-d₆300 MHz): δ 10.20 (s, 1H), 9.60 (s, 1H), 7.47 (d, J=3.0 Hz, 1H),7.24-7.18 (m, 6H), 7.01-6.96 (m, 3H), 6.79 (d, J=8.1 Hz, 2H), 4.06 (q,J=6.0 Hz, 2H), 3.96 (d, J=5.1 Hz, 1H), 3.14-3.07 (m, 2H), 1.31 (t, J=7.2Hz, 3H); MS (ESI): m/z 430.9 [M+H⁺].

Example 2 Compound 2,4-(3-ethoxy-4-hydroxy-5-nitrophenyl)-6-(pyridin-3-yl)-5-(thiophen-3-yl)-3,4-dihydropyridin-2(1H)-one

A mixture of 1-(pyridin-3-yl)-2-(thiophen-3-yl)ethanone (Intermediate 1,see Example 18 for synthesic description of all intermediates) (230 mg,1.1 mmol), 3-ethoxy-4-hydroxy-5-nitrobenzaldehyde (296 mg, 1.4 mmol),meldrum's acid (202 mg, 1.4 mmol) and AcONH₄ (108 mg, 1.4 mmol) in AcOH(3 mL) was refluxed under N₂ overnight. The mixture was concentratedunder reduced pressure and the residue was purified by prep-HPLC (0.1%TFA as additive) to give Compound 2 (60 mg, 12%). ¹H NMR (MeOD 400 MHz):δ 8.70 (d, J=5.2 Hz, 1H), 8.67 (s, 1H), 8.30 (d, J=8.0 Hz, 1H),7.90-7.62 (m, 1H), 7.72-7.60 (m, 1H), 7.27-7.24 (m, 2H), 6.94-6.92 (m,1H), 6.58 (d, J=4.8 Hz, 1H), 4.20-4.11 (m, 3H), 3.30-3.25 (m, 1H),2.79-2.74 (dd, J=16.2, 3.2 Hz, 1H), 1.45 (t, J=6.8 Hz, 3H). MS (ESI):m/z 438.3 [M+1]⁺.

Example 3 Compound 3,4-(4-(2H-tetrazol-5-yl)phenyl)-5-phenyl-6-(pyridin-3-yl)-3,4-dihydropyridin-2(1H)-one

A mixture of 2-phenyl-1-(pyridin-3-yl)ethanone (Intermediate 2) (100 mg,0.50 mmol), 4-(2H-tetrazol-5-yl)benzaldehyde (500 mg), meldrum's acid(88 mg, 0.61 mmol) and NH₄OAc (47 mg, 0.61 mmol) in AcOH (5 mL) wasrefluxed under N₂ overnight. The mixture was concentrated under reducedpressure and purified by prep-HPLC (0.1% TFA as additive) to giveCompound 3 (25 mg, yield 13%) as yellow solid. ¹H NMR (CD₃OD 400 MHz): δ8.68-8.50 (m, 2H), 8.21-8.15 (m, 1H), 8.10-8.00 (m, 2H), 7.75-7.60 (m,3H), 7.20-7.05 (m, 3H), 7.03-6.88 (m, 2H), 4.31-4.20 (m, 1H), 3.45-3.30(m, 1H), 2.83-2.71 (m, 1H). MS (ESI): m/z 395.4 [M+1]⁺.

Example 4 Compound 4,4-(3-ethoxy-4-hydroxy-5-nitrophenyl)-6-(1-methyl-1H-pyrazol-4-yl)-5-(thiophen-3-yl)-3,4-dihydropyridin-2(1H)-one

A mixture of 1-(1-methyl-1H-pyrazol-4-yl)-2-(thiophen-3-yl)ethanone(Intermediate 3) (150 mg, 0.73 mmol),3-ethoxy-4-hydroxy-5-nitrobenzaldehyde (185 mg, 0.87 mmol), meldrum'sacid (125 mg, 0.87 mmol) and AcONH₄ (84 mg, 1.1 mmol) in AcOH (3 mL) wasrefluxed under N₂ overnight. The mixture was concentrated under reducedpressure and the residue was purified by prep-HPLC (0.1% TFA asadditive) to give Compound 4 as yellow solid (20 mg, 6%). ¹H NMR(DMSO-d6, 400 MHz): δ 10.20 (s, 1H), 9.43 (s, 1H), 7.62 (s, 1H),7.42-7.36 (m, 2H), 7.19-7.11 (m, 1H), 7.06 (s, 2H), 6.70-6.68 (m, 1H),4.11-4.03 (m, 2H), 3.95-3.92 (m, 1H), 3.77 (s, 3H), 3.08 (dd, J=16.4,7.6 Hz, 1H), 2.60-2.52 (m, 1H), 1.34 (t, J=6.8 Hz, 3H). MS (ESI): m/z440.9 [M+1]⁺.

Example 5 Compound 5,4-(4-(2H-tetrazol-5-yl)phenyl)-6-(1-methyl-1H-pyrazol-4-yl)-5-(thiophen-3-yl)-3,4-dihydropyridin-2(1H)-one

A mixture of 1-(1-methyl-1H-pyrazol-4-yl)-2-phenylethanone (Intermediate4) (400 mg, 2.0 mmol), 4-(2H-tetrazol-5-yl)benzaldehyde (420 mg, 2.4mmol), meldrum's acid (345 mg, 2.4 mmol) and AcONH₄ (185 mg, 2.4 mmol)in AcOH (4 mL) was refluxed under N₂ overnight. The mixture wasconcentrated in vacuo and the residue was purified by prep-HPLC (0.1%TFA as additive) to give Compound 5 (25 mg, 3%). ¹H NMR (DMSO-d6 400MHz): δ 9.46 (brs, 1H), 7.98 (d, J=8.0 Hz, 2H), 7.58-7.48 (m, 3H),7.25-7.16 (m, 3H), 7.15 (d, J=6.4 Hz, 2H), 6.91 (s, 1H), 4.00 (d, J=5.2Hz, 1H), 3.72 (s, 3H), 3.20 (dd, J=16.0 Hz, 7.6 Hz, 1H), 2.50-2.45 (m,1H). MS (ESI): m/z 398.0 [M+1]⁺.

Example 6 Compound 6,2-hydroxy-4-(2-oxo-5,6-diphenyl-1,2,3,4-tetrahydropyridin-4-yl)benzoicacid

A mixture of 1,2-diphenylethanone (800 mg, 4.0 mmol), meldrum's acid(692 mg, 4.8 mmol), methyl 4-formyl-2-hydroxybenzoate (Intermediate 5)(742 mg, 4.0 mmol) and NH₄OAc (370 mg, 4.8 mmol) in AcOH (10 mL) wasrefluxed under N₂ overnight. The mixture was concentrated under reducedpressure and the residue was taken up in MeOH (10 mL) and aq. NaOH (2 M,10 mL) and was stirred at 60° C. under N₂ for 6 hours. The resultingmixture was cooled to room temperature, and then acidified with aqueousHCl (2 M) to pH=4, extracted with ethyl acetate (50 mL×3). The combinedorganic layers were dried over anhydrous Na₂SO₄ and concentrated underreduced pressure. The residue was purified by prep-HPLC (0.1% TFA asadditive) to give Compound 6 (195 mg, 2-step yield 12%) as off-whitesolid. ¹H NMR (DMSO-d6 400 MHz): δ 11.29 (brs, 1H), 9.63 (brs, 1H), 7.79(d, J=8.4 Hz, 1H), 7.36-7.22 (m, 5H), 7.10-6.98 (m, 5H), 6.86-6.79 (m,2H), 4.00 (d, J=6.0 Hz, 1H), 3.21 (dd, J=16.0, 7.6 Hz, 1H), 2.49-2.41(m, 1H). MS (ESI): m/z 385.7 [M+1]⁺.

Example 7 Compound 7,2-hydroxy-4-(2-oxo-6-phenyl-5-(thiophen-3-yl)-1,2,3,4-tetrahydropyridin-4-yl)benzoicacid

To a mixture of 1-phenyl-2-(thiophen-3-yl)ethanone (Intermediate 6) (300mg, 1.5 mmol), methyl 4-formyl-2-hydroxybenzoate (Intermediate 5) (325mg, 1.8 mmol), meldrum's acid (260 mg, 1.8 mmol) and AcONH₄ (140 mg, 1.8mmol) in AcOH (3 mL) was refluxed overnight. The mixture wasconcentrated in vacuo and the residue was taken up in MeOH (10 mL) andaq. NaOH (2 M, 10 mL) and was stirred at 40° C. overnight. The resultingmixture was cooled to room temperature, and then acidified with aq. HCl(2 M, 12 mL) to pH=5. Followed a standard aqueous/EtOAc workup i.e. themixture was extracted with EtOAc (3 times), washed with brine, thencombined organic layers were dried over anhydrous Na₂SO₄ andconcentrated in vacuo. The residue was purified by prep-HPLC (0.1% TFAas additive) to give Compound 7 (25 mg, 2-steps yield 4%) as yellowsolid. ¹H NMR (DMSO-d6 400 MHz): δ 9.60 (s, 1H), 7.78 (d, J=8.0 Hz, 1H),7.44-7.36 (m, 3H), 7.35-7.26 (m, 3H), 7.22-7.15 (m, 1H), 7.14-6.98 (m,2H), 6.82-6.74 (m, 1H), 6.25 (d, J=4.8 Hz, 1H), 4.13-4.05 (m, 1H), 3.18(dd, J=16.2, 8.0 Hz, 1H), 2.46-2.42 (m, 1H). MS (ESI): m/z 391.7 [M+1]⁺.

Example 8 Compound 8,4-(4-(2H-tetrazol-5-yl)phenyl)-6-(1-methyl-1H-pyrazol-4-yl)-5-phenyl-3,4-dihydropyridin-2(1H)-one

A mixture of 1-(1-methyl-1H-pyrazol-4-yl)-2-phenylethanone (Intermediate4) (400 mg, 2.0 mmol), 4-(2H-tetrazol-5-yl)benzaldehyde (420 mg, 2.4mmol), meldrum's acid (345 mg, 2.4 mmol) and AcONH₄ (185 mg, 2.4 mmol)in AcOH (4 mL) was refluxed under N₂ overnight. The mixture wasconcentrated in vacuo and the residue was purified by prep-HPLC (0.1%TFA as additive) to give Compound 8 (25 mg, 3%). ¹H NMR (DMSO-d6 400MHz): δ 9.46 (brs, 1H), 7.98 (d, J=8.0 Hz, 2H), 7.58-7.48 (m, 3H),7.25-7.16 (m, 3H), 7.15 (d, J=6.4 Hz, 2H), 6.91 (s, 1H), 4.00 (d, J=5.2Hz, 1H), 3.72 (s, 3H), 3.20 (dd, J=16.0 Hz, 7.6 Hz, 1H), 2.50-2.45 (m,1H). MS (ESI): m/z 398.0 [M+1]⁺.

Example 9 Compound 9,2-hydroxy-4-(6-(1-methyl-1H-pyrazol-4-yl)-2-oxo-5-(thiophen-3-yl)-1,2,3,4-tetrahydropyridin-4-yl)benzoicacid

A mixture of 1-(1-methyl-1H-pyrazol-4-yl)-2-(thiophen-3-yl)ethanone(Intermediate 3) (300 mg, 1.5 mmol), methyl 4-formyl-2-hydroxybenzoate(Intermediate 5) (315 mg, 1.7 mmol), meldrum's acid (245 mg, 1.7 mmol)and AcONH₄ (130 mg, 1.7 mmol) in AcOH (3 mL) was refluxed under N₂atmosphere overnight. The mixture was concentrated under reducedpressure and taken up in MeOH (10 mL) and aq. NaOH (2 M, 10 mL). Thereaction was stirred at 40° C. for 5 hours. The resulting mixture wascooled to room temperature, and then acidified with aq. HCl (2 M, 12 mL)to pH=5, followed by a standard aqueous/EtOAc workup. The residue waspurified by prep-HPLC (0.1% TFA as additive) to give Compound 9 as agray solid (45 mg, 2-steps yield 8%). ¹H NMR (DMSO-d6 400 MHz): δ 11.24(brs, 1H), 9.41 (brs, 1H), 7.73 (d, J=8.4 Hz, 1H), 7.63 (s, 1H), 7.38(dd, J=4.8, 2.8 Hz, 1H), 7.08 (s, 1H), 7.03 (d, J=1.6 Hz, 1H), 6.90-6.82(m, 2H), 6.67 (d, J=5.2 Hz, 1H), 3.94 (d, J=6.4 Hz, 1H), 3.77 (s, 3H),3.12 (dd, J=16.0, 7.6 Hz, 1H), 2.48-2.38 (m, 1H). MS (ESI): m/z 395.9[M+1]⁺.

Example 10 Compound 10,2-fluoro-6-hydroxy-4-(2-oxo-5,6-diphenyl-1,2,3,4-tetrahydropyridin-4-yl)benzoicacid

A mixture of 1, 2-diphenyl-ethanone (250 mg, 1.3 mmol), methyl2-fluoro-4-formyl-6-hydroxybenzoate (Intermediate 7) (252 mg, 1.3 mmol),meldrum's acid (230 mg, 1.6 mmol) and NH₄OAc (125 mg, 1.6 mmol) in AcOH(5 mL) was refluxed under N₂ overnight. The mixture was concentrated invacuo and the residue was taken up in MeOH (5 mL) and aq. NaOH (2 M, 5mL). This was stirred at 60° C. under N₂ for 5 hours. The resultingmixture was cooled to room temperature, and then acidified with aqueousHCl (2 M) to pH=4, followed by a standard aqueous/EtOAc workup. Theresidue was purified by prep-HPLC (0.1% TFA as additive) to giveCompound 10 (28 mg, 2-step yield 5%) as gray solid. ¹H NMR (DMSO-d6 400MHz): δ 9.64 (brs, 1H), 7.24-7.21 (m, 5H), 7.12-6.98 (m, 3H), 6.88-6.78(m, 3H), 6.74 (d, J=11.2 Hz, 1H), 4.01-3.90 (m, 1H), 3.20 (dd, J=16.4,7.6 Hz, 1H), 2.49-2.41 (m, 1H). MS (ESI): m/z 403.9 [M+1]⁺.

Example 11 Compound 11,2-fluoro-6-hydroxy-4-(6-(1-methyl-1H-pyrazol-4-yl)-2-oxo-5-(thiophen-3-yl)-1,2,3,4-tetrahydropyridin-4-yl)benzoicacid

A mixture of 1-(1-methyl-1H-pyrazol-4-yl)-2-(thiophen-3-yl)ethanone(Intermediate 3) (250 mg, 1.2 mmol), methyl2-fluoro-4-formyl-6-hydroxybenzoate (Intermediate 7) (240 mg, 1.2 mmol),meldrum's acid (217 mg, 1.5 mmol) and NH₄OAc (116 mg, 1.5 mmol) in AcOH(5 mL) was refluxed under N₂ overnight. The mixture was concentratedunder reduced pressure and the residue was taken up in MeOH (5 mL) and2M NaOH (5 mL). The reaction was stirred at 60° C. under N₂ for 5 hours.After being cooled to room temperature, the mixture was acidified withaqueous HCl (2 M) to pH=4 followed by a standard aqueous/EtOAc workup.The residue was purified by prep-HPLC (0.1% TFA as additive) to giveCompound 11 (14 mg, 2-step yield 2%) as gray solid. ¹H NMR (DMSO-d₆ 400MHz): δ 9.42 (brs, 1H), 7.63 (s, 1H), 7.39 (dd, J=4.8, 3.2 Hz, 1H), 7.10(s, 1H), 7.06 (d, J=2.8 Hz, 1H), 6.75-6.55 (m, 3H), 3.90 (d, J=6.4 Hz,1H), 3.77 (s, 3H), 3.10 (dd, J=16.0, 7.6 Hz, 1H), 2.49-2.38 (m, 1H). MS(ESI): m/z 413.8 [M+1]⁺.

Example 12 Compound 12,2-ethoxy-6-hydroxy-4-(2-oxo-5,6-diphenyl-1,2,3,4-tetrahydropyridin-4-yl)benzoicacid

A mixture of 1,2-diphenylethanone (216 mg, 1.1 mmol), methyl2-ethoxy-4-formyl-6-hydroxybenzoate (Intermediate 8) (304 mg, 1.3 mmol),meldrum's acid (260 mg, 1.8 mmol) and NH₄OAc (140 mg, 1.8 mmol) in HOAc(5 mL) was refluxed under N₂ atmosphere overnight. The mixture wasconcentrated under reduced pressure and the resulting residue was takenup in MeOH (10 mL) and aq. NaOH (2 M, 10 mL). The reaction was stirredat 70° C. under N₂ overnight. The resulting mixture was cooled to roomtemperature, and then acidified with aqueous HCl (2 M) to pH=3, followedby a standard aqueous/EtOAc workup. The residue was purified byprep-HPLC (0.1% TFA as additive) to give Compound 12 (22 mg, 2-stepyield 5%) as off-white solid. ¹H NMR (CD₃OD 400 MHz): δ 7.36-7.26 (m,5H), 7.10-7.03 (m, 3H), 6.96-6.90 (m, 2H), 6.73 (d, J=0.8 Hz, 1H), 6.67(s, 1H), 4.24 (q, J=7.2 Hz, 2H), 4.02 (dd, J=7.6, 2.4 Hz, 1H), 3.35-3.25(m, 1H), 2.68 (dd, J=16.0, 2.8 Hz, 1H), 1.46 (t, J=7.2 Hz, 3H). MS(ESI): m/z 429.8 [M+1]⁺.

Example 13 Compound 13,4-(6-cyclohexyl-2-oxo-5-(thiophen-3-yl)-1,2,3,4-tetrahydropyridin-4-yl)-2-fluoro-6-hydroxybenzoicacid

A mixture of 1-cyclohexyl-2-(thiophen-3-yl)ethanone (Intermediate 9)(271 mg, 1.3 mmol), methyl 2-fluoro-4-formyl-6-hydroxybenzoate(Intermediate 7) (300 mg, 1.5 mmol), meldrum's acid (216 mg, 1.5 mmol)and NH₄OAc (116 mg, 1.5 mmol) in HOAc (5 mL) was refluxed under N₂overnight. The mixture was concentrated under reduced pressure and theresidue was taken up in MeOH (10 mL) and aq. NaOH (2 M, 10 mL). Thereaction was stirred at 50° C. under N₂ overnight. The resulting mixturewas cooled, and then acidified with aqueous HCl (2 M) to pH=3, followedby a standard aqueous/EtOAc workup. The residue was purified byprep-HPLC (0.1% TFA as additive) to give Compound 13 (48 mg, 2-stepyield 9%) as off-white solid. ¹H NMR (CD₃OD 400 MHz): δ 7.38 (dd, J=5.2Hz, 2.8 Hz, 1H), 7.08 (d, J=2.4 Hz, 1H), 6.92 (d, J=3.2 Hz, 1H), 6.66(s, 1H), 6.53 (d, J=11.6 Hz, 1H), 3.84 (d, J=6.4 Hz, 1H), 3.11 (dd,J=16.4, 8.0 Hz, 1H), 2.76-2.66 (m, 1H), 2.54 (dd, J=16.4, 2.0 Hz, 1H),1.92-1.52 (m, 7H), 1.38-1.18 (m, 3H). MS (ESI): m/z 415.8 [M+1]⁺.

Example 14 Compound 14,4-(4-hydroxy-3-(2-hydroxyethoxy)-5-nitrophenyl)-5,6-diphenyl-3,4-dihydropyridin-2(1H)-one

A mixture of 1,2-diphenylethanone (300 mg, 1.5 mmol),4-hydroxy-3-(2-hydroxyethoxy)-5-nitrobenzaldehyde (350 mg, 1.5 mmol),meldrum's acid (220 mg, 1.8 mmol) and NH₄OAc (140 mg, 1.8 mmol) in AcOH(5 mL) was refluxed under N₂ overnight. The mixture was concentratedunder reduced pressure and the residue was taken up in MeOH (5 mL) andpotassium carbonate (400 mg, 3.0 mmol) was added. The reaction wasstirred at 25° C. for 2 hours. The mixture was poured into water,acidified with aqueous HCl (2 M) to pH=4, followed by a standardaqueous/EtOAc workup. The residue was purified by prep.-HPLC (0.1% TFAas additive) to give Compound 14 (36 mg, 2-step yield 6%) as yellowsolid. ¹H NMR (DMSO-d6 400 MHz): δ 9.62 (brs, 1H), 7.52 (s, 1H),7.36-7.17 (m, 6H), 7.10-6.96 (m, 3H), 6.85 (d, J=6.8 Hz, 2H), 4.10-4.04(m, 2H), 4.00 (d, J=5.6 Hz, 1H), 3.76 (t, J=4.8 Hz, 2H), 3.17 (dd,J=16.4, 7.2 Hz, 1H), 2.55 (m, 1H). MS (ESI): m/z 446.8 [M+1]⁺.

Example 15 Compound 15,2-fluoro-6-hydroxy-4-(1-methyl-2-oxo-5,6-diphenyl-1,2,3,4-tetrahydropyridin-4-yl)benzoicacid

Step 1: Synthesis of methyl2-fluoro-6-hydroxy-4-(2-oxo-5,6-diphenyl-1,2,3,4-tetrahydropyridin-4-yl)benzoate

A mixture of 1,2-diphenylethanone (300 mg, 1.5 mmol), methyl2-fluoro-4-formyl-6-hydroxybenzoate (Intermediate 7) (300 mg, 1.5 mmol),meldrum's acid (260 mg, 1.8 mmol) and NH₄OAc (140 mg, 1.8 mmol) in AcOH(5 mL) was refluxed under N₂ overnight. The mixture was concentratedunder reduced pressure and the residue was purified by silica gelchromatography (PE:EtOAc=5:1) to give product (90 mg, 16%) as a yellowsolid.

Step 2: Synthesis of methyl2-fluoro-6-(methoxymethoxy)-4-(2-oxo-5,6-diphenyl-1,2,3,4-tetrahydropyridin-4-yl)benzoate

To a mixture of the above product (90 mg, 0.22 mmol) and potassiumcarbonate (91 mg, 0.66 mmol) in DMF (5 mL) was addedchloro(methoxy)methane (35 mg, 0.44 mmol) at 25° C. and stirred for 2hours. Followed a standard aqueous/EtOAc workup to give product (90 mg,91%).

Step 3: Synthesis of methyl2-fluoro-6-(methoxymethoxy)-4-(1-methyl-2-oxo-5,6-diphenyl-1,2,3,4-tetrahydropyridin-4-yl)benzoate

To a mixture of the above product (90 mg, 0.20 mmol) and potassiumcarbonate (80 mg, 0.60 mmol) in DMF (5 mL) was added iodomethane (57 mg,0.40 mmol) at 25° C. and stirred for 2 hours. A standard aqueous/EtOAcworkup was followed to give product (90 mg, 97%), which was useddirectly to the next step without further purification.

Step 4: Synthesis of2-fluoro-6-hydroxy-4-(1-methyl-2-oxo-5,6-diphenyl-1,2,3,4-tetrahydropyridin-4-yl)benzoicacid

A mixture of the above product in MeOH (5 mL) and 2N NaOH (5 mL) wasstirred at 60° C. under N₂ atmosphere for 5 hours. After being cooled toroom temperature, the mixture was acidified with aqueous HCl (2 M) topH=4, followed by a standard aqueous/EtOAc workup. The residue wasdissolved in MeOH (10 mL) and conc. HCl (0.5 mL) was added. After beingstirred at 25° C. for 2 hours, the mixture was concentrated underreduced pressure. The residue was purified by prep.-HPLC (0.1% TFA asadditive) to give Compound 15 (25 mg, yield 29%) as white solid. ¹H NMR(DMSO-d₆ 400 MHz): δ 7.40-7.26 (m, 5H), 7.08-6.97 (m, 3H), 6.89 (s, 1H),6.80-6.74 (m, 3H), 3.85 (dd, J=6.4, 2.0 Hz, 1H), 3.28 (dd, J=16.0, 7.2Hz, 1H), 2.68 (s, 3H), 2.60 (dd, J=15.6, 2.4 Hz, 1H). MS (ESI): m/z417.7 [M+1]⁺.

Example 16 Compound 16,4-(3-ethoxy-4-hydroxy-5-nitrophenyl)-1-methyl-5,6-diphenyl-3,4-dihydropyridin-2(1H)-one

Step 1: Synthesis of4-(3-ethoxy-4-(methoxymethoxy)-5-nitrophenyl)-5,6-diphenyl-3,4-dihydropyridin-2(1H)-one

To a mixture of4-(3-ethoxy-4-hydroxy-5-nitrophenyl)-5,6-diphenyl-3,4-dihydropyridin-2(1H)-one(Compound 1, see Example 1) (200 mg, 0.47 mmol) and potassium carbonate(130 mg, 0.93 mmol) in DMF (5 mL) was added chloro(methoxy)methane (75mg, 0.93 mmol) at 25° C. and then the mixture was stirred at the sametemperature for 2 hours. The mixture was poured into water and astandard aqueous/EtOAc workup was followed to give product (200 mg,98%).

Step 2: Synthesis of4-(3-ethoxy-4-(methoxymethoxy)-5-nitrophenyl)-1-methyl-5,6-diphenyl-3,4-dihydropyridin-2(1H)-one

To a mixture of the above compound (200 mg, 0.42 mmol) and potassiumcarbonate (580 mg, 4.2 mmol) in DMF (5 mL) was added iodomethane (600mg, 4.2 mmol) at 25° C. and then the mixture was stirred for 2 hours.The mixture was poured into water and a standard aqueous/EtOAc workupwas followed to give product (200 mg, 97%).

Step 3: Synthesis of4-(3-ethoxy-4-hydroxy-5-nitrophenyl)-1-methyl-5,6-diphenyl-3,4-dihydropyridin-2(1H)-one

To a mixture of the above compound in MeOH (10 mL) was added conc. HCl(0.5 mL), then the mixture was stirred at 25° C. for 2 hours. Themixture was concentrated under reduced pressure. The residue waspurified by prep.-HPLC (0.1% TFA as additive) to give Compound 16 (120mg, yield 66%) as yellow solid. ¹H NMR (DMSO-d₆ 400 MHz): δ 7.52 (d,J=1.6 Hz, 1H), 7.48-7.39 (m, 3H), 7.38-7.24 (m, 3H), 7.06-6.98 (m, 3H),6.79 (d, J=6.4 Hz, 2H), 4.20-4.08 (m, 2H), 3.96-3.88 (m, 1H), 3.25 (dd,J=16.0, 6.4 Hz, 1H), 2.68 (s, 3H), 2.65 (d, J=4.0 Hz, 1H), 1.37 (t,J=7.2 Hz, 3H). MS (ESI): m/z 444.8 [M+1]⁺.

Example 17 Compound 17,2-fluoro-6-hydroxy-4-(6-(1-methyl-1H-pyrazol-4-yl)-2-oxo-5-(thiophen-2-yl)-1,2,3,4-tetrahydropyridin-4-yl)benzoicacid

A mixture of 1-(1-methyl-1H-pyrazol-4-yl)-2-(thiophen-2-yl)ethanone(Intermediate 12) (250 mg, 1.20 mmol),2-fluoro-4-formyl-6-hydroxybenzoic acid (Intermediate 11), meldrum'sacid (202 mg, 1.40 mmol) and NH₄OAc (108 mg, 1.40 mmol) in AcOH (5 mL)was refluxed under N₂ overnight. The reaction was poured into water anda standard aqueous/EtOAc workup was followed. The residue was purifiedby prep. HPLC (0.1% TFA as additive) to give compound 17 (17 mg, yield3%). ¹H NMR (CD₃OD 400 MHz): δ 7.62 (s, 1H), 7.33 (s, 1H), 7.24 (dd,J=5.2, 1.2 Hz, 1H), 6.90 (dd, J=5.2, 3.6 Hz, 1H), 6.79 (s, 1H), 6.77(dd, J=3.6, 0.8 Hz, 1H), 6.66 (dd, J=11.6, 1.6 Hz, 1H), 4.05 (dd, J=8.0,2.0 Hz, 1H), 3.89 (s, 3H), 3.25 (dd, J=16.4, 8.0 Hz, 1H), 2.64 (dd,J=16.4, 2.0 Hz, 1H). MS (ESI): m/z 413.7 [M+1]⁺.

Example 18 Synthesis of Intermediates Intermediate 1:1-(pyridin-3-yl)-2-(thiophen-3-yl)ethanone Step 1: Synthesis ofnicotinoyl chloride

To a solution of nicotinic acid (2 g, 16.2 mmol) in anhydrous THF (30mL) was added SOCl₂ (2.4 mL, 32.5 mmol). After stirring at 80° C. for 2hours, the mixture was concentrated in vacuo

Step 2: Synthesis of ethyl3-oxo-3-(pyridin-3-yl)-2-(thiophen-3-yl)propanoate

To a solution of ethyl 2-(thiophen-3-yl)acetate (2.3 g, 16.2 mmol) inanhydrous THF (20 mL) was added LiHMDS (19.4 mL, 19.44 mmol) at −78° C.After stirring at that temperature for 0.5 h, a solution of nicotinoylchloride (2.8 g, 16.2 mmol) in anhydrous THF (10 mL) was added into thereaction mixture and stirred at −78° C. for 4 hours. The mixture wasquenched with NH₄Cl solution, extracted with EtOAc. The organic layerwas concentrated in vacuo and purified by silica gel columnchromatography (1.7 g, yield 38%).

Step 3: Synthesis of 1-(pyridin-3-yl)-2-(thiophen-3-yl)ethanone

To a solution of ethyl3-oxo-3-(pyridin-3-yl)-2-(thiophen-3-yl)propanoate (2 g, 7.26 mmol) inDMSO (20 mL) was added catalytic amount of brine (0.2 mL). The reactionmixture was heated at 160° C. for 2 h. The reaction mixture was cooledto room temperature, diluted with water, and extracted with EtOAc. Theorganic layer was concentrated in vacuo and purified by columnchromatography to afford Intermediate 1 (1 g, yield 68%).

Intermediate 2: Synthesis of 2-phenyl-1-(pyridin-3-yl)ethanone Step 1

A mixture of nicotinic acid (2 g, 16.2 mmol), N,O-dimethylhydroxylaminehydrochloride (1.6 g, 16.2 mmol), EDCI (3.2 g, 16.2 mmol), HOBT (2.5 g,16.2 mmol), and Et₃N (6.7 mL, 48.7 mmol) in DCM (30 mL) was stirred atroom temperature for 4 h. The mixture was concentrated in vacuo andpurified by silica gel column chromatography (PE:EtOAc=2:1) to giveproduct (1.4 g, 52%).

Step 2

To a solution of the above product (Step 1) (500 mg, 3.0 mmol) inanhydrous THF (10 mL) was added benzylmagnesium chloride (2 M/L in THF,1.8 mL, 3.6 mmol) drop wise at −78° C. The reaction was stirred at −78°C. for 2 h, then quenched with aqueous NH₄Cl and extracted with EtOAc.The organic layer was dried over Na₂SO₄, concentrated in vacuo andpurified by column chromatography (PE:EtOAc=5:1) to give Intermediate 2(110 mg, 19%).

Intermediate 3: 1-(1-methyl-1H-pyrazol-4-yl)-2-(thiophen-3-yl)ethanone

Followed same three step procedure described in Intermediate 1.

Step 1: Synthesis of 1-methyl-1H-pyrazole-4-carbonyl chloride

Started with 1-methyl-1H-pyrazole-4-carboxylic acid, and used tolueneinstead of THF.

Step 2: Synthesis of ethyl3-(1-methyl-1H-pyrazol-4-yl)-3-oxo-2-(thiophen-3-yl)propanoate

Used crude from step 1 and ethyl 2-(thiophen-3-yl)acetate. Afterstirring at −78° C., the reaction was warmed to room temperature andstirred overnight.

Step 3: Synthesis of1-(1-methyl-1H-pyrazol-4-yl)-2-(thiophen-3-yl)ethanone

The reaction mixture was stirred at 180° C. for 2 hours. Purified bysilica gel column chromatography (PE:EtOAc=10:1) to give Intermediate 3(160 mg, yield 61.8%).

Intermediate 4: 1-(1-methyl-1H-pyrazol-4-yl)-2-phenylethanone Step 1:Synthesis of N-methoxy-N,1-dimethyl-1H-pyrazole-4-carboxamide

A mixture of 1-methyl-1H-pyrazole-4-carboxylic acid (5.0 g, 39.6 mmol),N,O-dimethylhydroxylamine hydrochloride (4.8 g, 48.0 mmol),1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (9.2 g, 48.0mmol), hydroxybenzotriazole (6.5 g, 48.0 mmol) and triethylamine (12.1g, 120 mmol) in dichloromethane (100 mL) was stirred at 9-17° C.overnight. The reaction mixture was diluted with water (50 mL) and theresulting mixture was extracted with EtOAc (30 mL×3), the combinedorganic layers were washed with brine (50 mL) and dried over anhydrousNa₂SO₄, filtered and concentrated in vacuo (an example of a standardaqueous/EtOAc procedure), then purified by silica gel columnchromatography (PE:EtOAc=5:1) to give product (4.6 g, yield 69%).

Step 2: Synthesis of 1-(1-methyl-1H-pyrazol-4-yl)-2-phenylethanone

To a solution of N-methoxy-N,1-dimethyl-1H-pyrazole-4-carboxamide (4.6g, 27.2 mmol) in anhydrous THF (50 mL) was added benzylmagnesiumchloride (2.0 M in THF, 16.7 mL, 32.6 mmol) at −78° C. under N₂, thenstirred at the same temperature for 3 hours. The mixture was quenchedwith saturated NH₄Cl solution, followed by a standard aqueous/EtOAcprocedure. The crude was purified by silica gel column chromatography(PE:EtOAc=10:1) to give Intermediate 4 (4.0 g, yield 74%).

Intermediate 5: methyl 4-formyl-2-hydroxybenzoate Step 1: Synthesis of4-formyl-2-methoxyphenyl trifluoromethanesulfonate

To a mixture of 4-hydroxy-3-methoxybenzaldehyde (10.0 g, 65.7 mmol) andEt₃N (13.0 g, 128.7 mmol) in anhydrous DCM (100 mL) was added Tf₂O (28.0g, 99.2 mmol) dropwise at room temperature and the reaction was stirredfor 1 hour. The resulting mixture was quenched with water (100 mL)followed by a standard aqueous/EtOAc workup. The residue was purified bysilica gel column chromatography (PE:EtOAc=20:1) to give the product asa colorless oil (17.0 g, yield 91%).

Step 2: Synthesis of methyl 4-formyl-2-methoxybenzoate

A mixture of the product from Step 1 (5.0 g, 17.6 mmol), Pd(OAc)₂ (500mg, 2.2 mmol), dppf (560 mg, 1.0 mmol) and Et₃N (3 mL) in MeOH (30 mL)and DMF (3 mL) was stirred under CO atmosphere (0.5 MPa) at 80° C.overnight. The reaction was cooled to room temperature and filtered. Thefiltrate was concentrated in vacuo. The residue was purified by columnchromatography on silica gel (PE:EtOAc=15:1) to give product as acolorless solid (2.6 g, yield 76%).

Step 3: Synthesis of methyl 4-formyl-2-hydroxybenzoate

A mixture of product from Step 2 (2.6 g, 13.4 mmol) and AlCl₃ (3.6 g,26.8 mmol) in anhydrous DCM (50 mL) was refluxed for 3 minutes. Theresulting mixture was cooled and poured into water (100 mL), followed bya standard aqueous/DCM workup to give Intermediate 5 (2.2 g, yield 92%).

Intermediate 6: Synthesis of 1-phenyl-2-(thiophen-3-yl)ethanone Step 1:Synthesis of N-methoxy-N-methyl-2-(thiophen-3-yl)acetamide

A mixture of 2-(thiophen-3-yl)acetic acid (2.0 g, 14.1 mmol),O,N-dimethylhydroxylamine (1.68 g, 16.9 mmol), EDCI (2.95 g, 15.5 mmol),HOBT (2.15 g, 15.5 mmol), and TEA (3.7 mL, 31 mmol) in anhydrous DCM (50mL) was stirred at room temperature under nitrogen for two hours. Thereaction mixture was diluted with CH₂Cl₂, and the organic layer waswashed with aqueous HCl solution (0.5 mol/L, 30 mL×2), saturated NaHCO₃(30 mL×2) and brine(30 mL), dried over Na₂SO₄, filtered, andconcentrated to give the crudeN-methoxy-N-methyl-2-(thiophen-3-yl)acetamide (2.0 g, yield 76.6%).

Step 2: Synthesis of 1-phenyl-2-(thiophen-3-yl)ethanone (Intermediate 2)

To the solution of bromo-benzene (1.0 g, 6.36 mmol) in anhydrous THF (30mL) was added n-BuLi (5.9 mmol, 2.4 mL) at −78° C. under nitrogen, andthe mixture was stirred at −78° C. for further 20 min., a solution ofN-methoxy-N-methyl-2-(thiophen-3-yl)acetamide (1 g, 5.4 mmol) inanhydrous THF (10 mL) was added. The resulting mixture was stirred at−78° C. under nitrogen for about 30 min., poured into NH₄Cl aqueoussolution, and extracted with EtOAc. The organic layers were washed withbrine, dried over sodium sulfate, concentrated, and purified by columnchromatography (PE:EtOAc=20:1) to afford Intermediate 6 (600 mg, yield55.0%).

Intermediate 7: methyl 2-fluoro-4-formyl-6-hydroxybenzoate Step 1:Synthesis of 2-fluoro-4-formyl-6-methoxyphenyl trifluoromethanesulfonate

Followed the procedure described in Intermediate 5, Step 1 where Tf₂Owas added at 0° C. and reaction was stirred at 0° C. Yield afterpurification was 61%.

Step 2: Synthesis of methyl 2-fluoro-4-formyl-6-methoxybenzoate

A mixture of the compound from step 1 (1.1 g, 3.6 mmol), Pd(OAc)₂ (200mg, 0.89 mmol), dppf (200 mg, 0.36 mmol) and Et₃N (2 mL) in MeOH (50 mL)and DMF (2 mL) was stirred under CO atmosphere (50 psi) at 80° C.overnight. The resulting mixture was cooled and filtered. A standardaqueous/EtOAc workup was followed by purification by columnchromatography on silica gel (PE:EtOAc=15:1) to give product as anoff-white solid (280 mg, yield 36%).

Step 3: Synthesis of methyl 2-fluoro-4-formyl-6-hydroxybenzoate

Followed procedure described in Intermediate 5, step 3 starting withmethyl 2-fluoro-4-formyl-6-methoxybenzoate (280 mg, 1.3 mmol) and AlCl₃(350 mg, 2.6 mmol) where reaction was complete after refluxing 10 min.Isolated 250 mg of Intermediate 7, yield 96%.

Intermediate 8: methyl 2-ethoxy-4-formyl-6-hydroxybenzoate Step 1:Synthesis of ethyl 4-bromo-3,5-diethoxybenzoate

A mixture of 4-bromo-3,5-dihydroxybenzoic acid (2.0 g, 8.6 mmol), EtI(6.7 g, 43.0 mmol) and K₂CO₃ (5.9 g, 43.0 mmol) in DMF (20 mL) wasstirred at 50° C. overnight. Followed standard aqueous/EtOAc workup (2.5g, yield 93%).

Step 2: Synthesis of (4-bromo-3,5-diethoxyphenyl)methanol

To a solution of the above from step 1 (5.0 g, 15.8 mmol) in anhydrousTHF (50 mL) was added DIBAL-H (1 M in toluene, 80 mL, 80.0 mmol)dropwise at −78° C. under N₂ and the reaction was stirred at −78° C. for2 hours. The resulting mixture was quenched with sat. aq. NH₄Cl (50 mL),followed by a standard aqueous/EtOAc workup to give product (4.2 g,yield 98%).

Step 3: Synthesis of methyl 2,6-diethoxy-4-(hydroxymethyl)benzoate

A mixture of the above from Step 2 (3.0 g, 10.9 mmol), Pd(OAc)₂ (500 mg,2.2 mmol), dppf (500 mg, 0.90 mmol) and Et₃N (5 mL) in MeOH (50 mL) andDMF (10 mL) was stirred under CO atmosphere (5 MPa) at 120° C. for 3days. The resulting mixture was cooled to room temperature and filtered.The filtrate was diluted with ethyl acetate (200 mL) and washed withbrine (50 mL×3). The organic layer was dried over anhydrous Na₂SO₄, andconcentrated in vacuo. The residue was purified by column chromatographyon silica gel (PE:EtOAc=8:1) (1.0 g, yield 36%).

Step 4: Synthesis of methyl 2,6-diethoxy-4-formylbenzoate

A mixture of the above product from Step 3 (1.0 g, 3.9 mmol) and MnO₂(1.0 g, 11.5 mmol) in DCM (30 mL) was refluxed for 1 hour. The mixturewas filtered and the filtrate was concentrated in vacuo to give product(950 mg, yield 96%) as an off-white solid.

Step 5: Synthesis of methyl 2-ethoxy-4-formyl-6-hydroxybenzoate

A mixture of methyl 2,6-diethoxy-4-formylbenzoate (950 mg, 3.8 mmol) andAlCl₃ (1.0 g, 7.6 mmol) in anhydrous DCM (30 mL) was stirred at 10° C.for 5 minutes. The resulting mixture was poured into water (100 mL)followed by a standard aqueous/EtOAc workup. The residue was purified bycolumn chromatography on silica gel (PE:EtOAc=10:1) to give Intermediate8 as off-white solid (500 mg, yield 59%).

Intermediate 9:1-cyclohexyl-2-(thiophen-3-yl)ethanone Step 1: Synthesisof cyclohexanecarbonyl chloride

Followed the procedure described in Step 1 of Intermediate 1, startingwith cyclohexanecarboxylic acid.

Step 2: Synthesis of ethyl3-cyclohexyl-3-oxo-2-(thiophen-3-yl)propanoate

Followed the procedure described in Step 2 of Intermediate 1. Used crudefrom step 1 and ethyl 2-(thiophen-3-yl)acetate. Crude product was takenforward without purification.

Step 3: Synthesis of 1-cyclohexyl-2-(thiophen-3-yl)ethanone

Followed the procedure described in Step 3 of Intermediate 1 to giveIntermediate 9 (352 mg, 3-step yield 43%).

Intermediate 10: 4-hydroxy-3-(2-hydroxyethoxy)-5-nitrobenzaldehyde Step1: Synthesis of 3-(2-hydroxyethoxy)-4-methoxybenzaldehyde

Followed procedure described in Intermediate 8, step 1 starting from3-hydroxy-4-methoxybenzaldehyde and 2-bromoethanol with the reactionconditions of 90° C. overnight. Isolated 400 mg, yield 62.5%.

Step 2: Synthesis of 4-hydroxy-3-(2-hydroxyethoxyl)benzaldehyde

To the mixture the crude from step 1 (400 mg, 2.0 mmol) in anhydrous DCM(10 mL) was added aluminum chloride (542 mg, 4.0 mmol), and then themixture was refluxed for 2 days. To the mixture was added dilute HCl andfollowed by a standard aqueous/EtOH workup. The residue was purified byreverse-phase preparatory HPLC (130 mg, yield 35.2%).

Step 3: Synthesis of 4-hydroxy-3-(2-hydroxyethoxy)-5-nitrobenzaldehyde

To the solution of 4-hydroxy-3-(2-hydroxyethoxyl)benzaldehyde in aceticacid (5 mL) was added 65% nitric acid (34.6 mg, 0.55 mmol) at 0° C., andthe mixture was stirred at 0° C. 4 hours. The reaction mixture waspoured into ice water followed by a standard aqueous/EtOH workup.Isolated 110 mg, yield 88.7% of Intermediate 10.

Intermediate 11: 2-fluoro-4-formyl-6-hydroxybenzoic acid

A mixture of methyl 2-fluoro-4-formyl-6-hydroxybenzoate (Intermediate 7)(300 mg, 1.50 mmol) and aqueous NaOH (2 M, 10 mL) in MeOH (10 mL) wasstirred at 20° C. for 5 hours. The mixture was acidified with aqueousHCl until pH=2 followed by a standard aqueous/EtOH workup. The residuewas used for next step without further purification.

Intermediate 12: 1-(1-methyl-1H-pyrazol-4-yl)-2-(thiophen-2-yl)ethanone

Followed same three step procedure described in Intermediate 1.

Step 1: Synthesis of 1-methyl-1H-pyrazole-4-carbonyl chloride

See Intermediate 3, step 1.

Step 2: Synthesis of ethyl3-(1-methyl-1H-pyrazol-4-yl)-3-oxo-2-(thiophen-2-yl)propanoate

Used crude from step 1 and ethyl 2-(thiophen-2-yl)acetate. Crude productwas used in next step.

Step 3: Synthesis of1-(1-methyl-1H-pyrazol-4-yl)-2-(thiophen-3-yl)ethanone

Reaction mixture was stirred at 160° C. for 4 hours. Purified by columnchromatography (PE:EtOAc=3:1) to give Intermediate 12 (346 mg, 3-stepyield 42%) as brown oil.

Example 19 GSNOR Assays

Various compounds were tested in vitro for their ability to inhibitGSNOR activity. GSNOR inhibitor compounds in Examples 1-17 had an IC₅₀of about <1.0 μM. GSNOR inhibitor compounds in Examples 1-2, 4, 6-7,9-14, and 17 had an IC₅₀ of about less than 0.1 μM. GSNOR expression andpurification is described in Biochemistry 2000, 39, 10720-10729.

GSNOR Fermentation:

Pre-cultures were grown from stabs of a GSNOR glycerol stock in 2XYTmedia containing 100 ug/ml ampicillin after an overnight incubation at37° C. Cells were then added to fresh 2XYT (4 L) containing ampicillinand grown to an OD (A₆₀₀) of 0.6-0.9 at 37° C. before induction. GSNORexpression was induced with 0.1% arabinose in an overnight incubation at20° C.

GSNOR Purification:

E. coli cell paste was lysed by nitrogen cavitation and the clarifiedlysate purified by Ni affinity chromatography on an AKTA FPLC (AmershamPharmacia). The column was eluted in 20 mM Tris pH 8.0/250 mM NaCl witha 0-500 mM imidazole gradient. Eluted GSNOR fractions containing theSmt-GSNOR fusion were digested overnight with Ulp-1 at 4° C. to removethe affinity tag then re-run on the Ni column under the same conditions.GSNOR was recovered in the flowthrough fraction and for crystallographyis further purified by Q-Sepharose and Heparin flowthroughchromatography in 20 mM Tris pH 8.0, 1 mM DTT, 10 uM ZnSO₄.

GSNOR Assay:

GSNO and Enzyme/NADH Solutions are made up fresh each day. The Solutionsare filtered and allowed to warm to room temperature. GSNO Solution: 100mM NaPO4 (pH 7.4), 0.480 mM GSNO. 396 μL of GSNO Solution is added to acuvette followed by 8 μL of test compound in DMSO (or DMSO only for fullreaction control) and mixed with the pipette tip. Compounds to be testedare made up at a stock concentration of 10 mM in 100% DMSO. 2 foldserial dilutions are done in 100% DMSO. 8 μL of each dilution are addedto an assay so that the final concentration of DMSO in the assay is 1%.The concentrations of compounds tested range from 100 to 0.003 μM.Enzyme/NADH Solution: 100 mM NaPO4 (pH 7.4), 0.600 mM NADH, 1.0 μg/mLGSNO Reductase. 396 μL of the Enzyme/NADH Solution is added to thecuvette to start the reaction. The cuvette is placed in the Cary 3EUV/Visible Spectrophotometer and the change in 340 nm absorbance/min at25° C. is recorded for 3 minutes. The assays are done in triplicate foreach compound concentration. IC50's for each compound are calculatedusing the standard curve analysis in the Enzyme Kinetics Module ofSigmaPlot.

Final assay conditions: 100 mM NaPO4, pH 7.4, 0.240 mM GSNO, 0.300 mMNADH, 0.5 μg/mL GSNO Reductase and 1% DMSO. Final volume: 800μL/cuvette.

Example 20 Human NK3 Receptor Binding Assay at 10 μM

Evaluation of the affinity of test compounds for the human neurokininNK3 receptor from transfected CHO cells was performed using anantagonist radioligand binding assay. Antagonist activity was evaluatedat a concentration of 10 μM for a representative subset of testcompounds, and the results were expressed as the percent inhibition atthat concentration.

Materials and Methods:

Receptor binding assays were performed using crude membranes preparedfrom CHO cells expressing human NK3 receptor. Osanetant (SR142801), anon-peptide NK3 antagonist, was used as a ligand, and SB222200 was usedas a positive control reference compound. The assay was performed withcell membrane homogenates (24 μg protein) incubated for 120 min at 22°C. with 0.4 nM [³H]SR142801 in the absence or presence of the testcompound in a buffer containing 20 mM Hepes/NaOH (pH 7.4), 120 mM NaCl,1 mM MnCl₂, 0.01% bacitracin, 0.002% aprotinin and 0.1% BSA. Followingincubation, the samples were filtered rapidly under vacuum through glassfiber filters (GF/B, Packard) that had been presoaked with 0.3% PEI. Thefilters were rinsed several times with ice-cold 50 mM Tris/HCl using a96-sample cell harvester (Unifilter, Packard), dried, and counted forradioactivity in a scintillation counter (Topcount, Packard) using ascintillation cocktail (Microscint 0, Packard). Each test compound wasevaluated at a single concentration of 10 μM. The standard referencecompound, SB222200, was tested in each experiment. Nonspecific bindingwas determined in the presence of 10 μM SB222200. The specific ligandbinding to the receptor is defined as the difference between the totalbinding and the nonspecific binding. The results are expressed as apercent of control specific binding ((measured specific binding/controlspecific binding)×100) obtained in the presence of the test compounds.

Results: The compounds in the following Examples had about >50%inhibition at 10 μM: Examples 1, 2, and 4.

Example 21 Human NK3 Receptor Binding Assay at 1 uM

Evaluation of the affinity of compounds for the human NK3 receptor fromtransfected CHO cells was performed using an antagonist radioligandbinding assay. Antagonist activity was evaluated at a concentration of 1μM for a representative subset of test compounds, and the results wereexpressed as the percent inhibition at that concentration.

Materials and Methods:

Receptor binding assays were performed using crude membranes preparedfrom CHO cells expressing human NK3 receptor. Osanetant (SR142801), anon-peptide NK3 antagonist, was used as a ligand, and SB222200 was usedas a positive control reference compound. The assay was performed withcell membrane homogenates (24 μg protein) incubated for 120 min at 22°C. with 0.4 nM [³H]SR142801 in the absence or presence of the testcompound in a buffer containing 20 mM Hepes/NaOH (pH 7.4), 120 mM NaCl,1 mM MnCl₂, 0.01% bacitracin, 0.002% aprotinin and 0.1% BSA. Followingincubation, the samples were filtered rapidly under vacuum through glassfiber filters (GF/B, Packard) that had been presoaked with 0.3% PEI. Thefilters were rinsed several times with ice-cold 50 mM Tris/HCl using a96-sample cell harvester (Unifilter, Packard), dried, and counted forradioactivity in a scintillation counter (Topcount, Packard) using ascintillation cocktail (Microscint 0, Packard). Each test compound wasevaluated at a single concentration of 1 μM. The standard referencecompound, SB222200, was tested in each experiment. Nonspecific bindingwas determined in the presence of 10 μM SB222200. The specific ligandbinding to the receptor is defined as the difference between the totalbinding and the nonspecific binding. The results are expressed as apercent of control specific binding ((measured specific binding/controlspecific binding)×100) obtained in the presence of the test compounds.

Results: The compounds in the following Examples had about >50%inhibition at 1 μM: Examples 14 and 16.

Example 22 Human NK3 Receptor Binding Assay Determined by IC₅₀

Evaluation of the affinity of compounds for the human NK3 receptor fromtransfected CHO cells was performed using an antagonist radioligandbinding assay. The antagonist activity of a representative subset oftest compounds was evaluated over a range of concentrations, and theresults were expressed as IC₅₀ values.

Materials and Methods:

Receptor binding assays were performed using crude membranes preparedfrom CHO cells expressing human NK3 receptor. Osanetant (SR142801), anon-peptide NK3 antagonist, was used as a ligand, and SB222200 was usedas a positive control reference compound. The assay was performed withcell membrane homogenates (24 μg protein) incubated for 120 min at 22°C. with 0.4 nM [³H]SR142801 in the absence or presence of the testcompound in a buffer containing 20 mM Hepes/NaOH (pH 7.4), 120 mM NaCl,1 mM MnCl₂, 0.01% bacitracin, 0.002% aprotinin and 0.1% BSA. Followingincubation, the samples were filtered rapidly under vacuum through glassfiber filters (GF/B, Packard) that had been presoaked with 0.3% PEI. Thefilters were rinsed several times with ice-cold 50 mM Tris/HCl using a96-sample cell harvester (Unifilter, Packard), dried, and counted forradioactivity in a scintillation counter (Topcount, Packard) using ascintillation cocktail (Microscint 0, Packard). For IC₅₀ generation,test compounds were assayed at 8 concentrations within the range of1×10⁻⁵ to 1×10⁻¹⁰ M (10 μM to 100 μM). The standard reference compound,SB222200, was tested in each experiment. Nonspecific binding wasdetermined in the presence of 10 μM SB222200. The specific ligandbinding to the receptor is defined as the difference between the totalbinding and the nonspecific binding. The IC₅₀ values (concentrationcausing a half-maximal inhibition of control specific binding) and Hillcoefficients (nH) were determined by non-linear regression analysis ofthe competition curves generated with mean replicate values using Hillequation curve fitting (Y=D+[(A−D)/(1+(C/C₅₀)^(nH))], where Y=specificbinding, D=minimum specific binding, A=maximum specific binding,C=compound concentration, C₅₀=IC₅₀, and nH=slope factor). This analysiswas performed using software developed at Cerep (Hill software) andvalidated by comparison with data generated by the commercial softwareSigmaPlot® 4.0 for Windows® (© 1997 by SPSS Inc.). The inhibitionconstants (K_(i)) were calculated using the Cheng Prusoff equation(K_(i)=IC₅₀/(1+(L/K_(D))), where L=concentration of radioligand in theassay, and K_(D)=affinity of the radioligand for the receptor). Ascatchard plot was used to determine the Kd.

Results: Compound 1 in Example 1 had an IC₅₀ of about 1.1 μM.

Example 23 Efficacy of GSNORi in Experimental Asthma

Experimental Asthma Model:

A mouse model of ovalbumin (OVA)-induced asthma is used to screen GSNORinhibitors for efficacy against methacholine (MCh)-inducedbronchoconstriction/airway hyper-reactivity. This is a widely used andwell characterized model that presents with an acute, allergic asthmaphenotype with similarities to human asthma. Efficacy of GSNORinhibitors are assessed using a prophylactic protocol in which GSNORinhibitors are administered prior to challenge with MCh.Bronchoconstriction in response to challenge with increasing doses ofMCh is assessed using whole body plethysmography (P_(enh); Buxco). Theamount of eosinophil infiltrate into the bronchoaveolar lavage fluid(BALF) is also determined as a measure of lung inflammation. The effectof GSNOR inhibitors are compared to vehicles and to Combivent (inhaled;IH) as the positive control

Materials and Methods

Allergen Sensitization and Challenge Protocol

OVA (500 μg/ml) in PBS is mixed with equal volumes of 10% (w/v) aluminumpotassium sulfate in distilled water and incubated for 60 min. at roomtemperature after adjustment to pH 6.5 using 10 N NaOH. Aftercentrifugation at 750×g for 5 min, the OVA/alum pellet is resuspended tothe original volume in distilled water. Mice receive an intraperitoneal(IP) injection of 100 μg OVA (0.2 mL of 500 μg/mL in normal saline)complexed with alum on day 0. Mice are anesthetized by IP injection of a0.2-mL mixture of ketamine and xylazine (0.44 and 6.3 mg/mL,respectively) in normal saline and are placed on a board in the supineposition. Two hundred fifty micrograms (100 μl of a 2.5 mg/ml) of OVA(on day 8) and 125 μg (50 μl of 2.5 mg/ml) OVA (on days 15, 18, and 21)are placed on the back of the tongue of each animal.

Pulmonary Function Testing (Penh)

In vivo airway responsiveness to methacholine is measured 24 h after thelast OVA challenge in conscious, freely moving, spontaneously breathingmice with whole body plethysmography using a Buxco chamber (Wilmington,N.C.). Mice are challenged with aerosolized saline or increasing dosesof methacholine (5, 20 and 50 mg/mL) generated by an ultrasonicnebulizer for 2 min. The degree of bronchoconstriction is expressed asenhanced pause (P_(enh)), a calculated dimensionless value, whichcorrelates with the measurement of airway resistance, impedance, andintrapleural pressure in the same mouse. P_(enh) readings are taken andaveraged for 4 min. after each nebulization challenge. P_(enh) iscalculated as follows: P_(enh)=[(T_(e)/T_(r)−1)×(PEF/PIF)], where T_(e)is expiration time, T_(r) is relaxation time, PEF is peak expiratoryflow, and PIF is peak inspiratory flow×0.67 coefficient. The time forthe box pressure to change from a maximum to a user-defined percentageof the maximum represents the relaxation time. The T_(r) measurementbegins at the maximum box pressure and ends at 40%.

Eosinophil Infiltrate in BALF

After measurement of airway hyper-reactivity, the mice are exsanguinatedby cardiac puncture, and then BALF is collected from either both lungsor from the right lung after tying off the left lung at the mainstembronchus. Total BALF cells are counted from a 0.05 mL aliquot, and theremaining fluid is centrifuged at 200×g for 10 min at 4° C. Cell pelletsare resuspended in saline containing 10% BSA with smears made on glassslides. Eosinophils are stained for 5 min. with 0.05% aqueous eosin and5% acetone in distilled water, rinsed with distilled water, andcounterstained with 0.07% methylene blue.

GSNOR Inhibitors and Controls

GSNOR inhibitors are reconstituted in phosphate buffered saline (PBS),pH 7.4, at concentrations ranging from 0.00005 to 3 mg/mL. GSNORinhibitors are administered to mice (10 mL/kg) as a single dose eitherintravenously (IV) or orally via gavage. Dosing is performed from 30min. to 24 h prior to MCh challenge. Effect of GSNOR inhibitors arecompared to PBS vehicle dosed in the same manner.

Combivent is used as the positive control in all studies. Combivent(Boehringer Ingelheim) is administered to the lung using the inhalerdevice supplied with the product, but adapted for administration tomice, using a pipet tip. Combivent is administered 48 h, 24 h, and 1 hprior to MCh challenge. Each puff (or dose) of Combivent provides a doseof 18 μg ipatropium bromide (IpBr) and 103 μg albuterol sulfate orapproximately 0.9 mg/kg IpBr and 5 mg/kg albuterol.

Statistical Analyses

Area under the curve values for P_(enh) across baseline, saline, andincreasing doses of MCh challenge are calculated using GraphPad Prism5.0 (San Diego, Calif.) and expressed as a percent of the respective (IVor orally administered) vehicle control. Statistical differences amongtreatment groups and the respective vehicle control group within eachstudy are calculated using one-way ANOVA, Dunnetts (JMP 8.0, SASInstitute, Cary, N.C.). A p value of <0.05 among the treatment groupsand the respective vehicle control group is considered significantlydifferent.

Example 24 Mouse Pharmacokinetic (PK) Study

Experimental Model

The mouse is used to determine the pharmacokinetics of compounds of theinvention. This species is widely used to assess the bioavailability ofcompounds by administering both oral (PO) and intravenous (IV) testarticles. Efficacy of the compounds of the invention are compared byassessing plasma exposure in male BALB/c mice either via IV or POadministration at the times of peak activity.

Materials and Methods

IV Administration of Compounds of the Invention

Compounds of the invention are reconstituted in a phosphate bufferedsaline (PBS)/10% Solutol (HS 15) clear solution resulting in aconcentration of 0.2 mg/mL and administered to mice (2 mg/kg) as asingle IV dose. Animals dosed via the lateral tail vein. Blood samplesare collected at designated time points (0.083, 0.25, 0.5, 1, 2, 4, 8,16, 24 hours) by cardiac puncture under isoflurane anesthesia (up to 1mL blood per animal). The blood is collected into tubes containingLi-Heparin. The blood samples are kept on ice until centrifugationwithin approximately 30 minutes of collection. The plasma is transferredinto labeled polypropylene tubes and frozen at −70° C. until analyzed byLC/MS/MS.

PO Administration of Compounds of the Invention

The compounds of the invention are reconstituted in 40% PropyleneGlycol/40% Propylene Carbonate/20% of a 5% Sucrose clear solutionresulting in a concentration of 2 mg/mL and administered to mice (10mg/kg) as a single oral dose via gavage. Blood samples are collected at0.25, 0.5, 1, 2, 4, 8, 12, 16, 20 and 24 hours post dose by cardiacpuncture under isoflurane anesthesia. The blood is collected in tubescontaining Li-Heparin. The blood samples are kept on ice untilcentrifugation within approximately 30 minutes of collection. The plasmais transferred into labeled polypropylene tubes and frozen at −70° C.until analyzed by LC/MS/MS.

LC/MS/MS Analysis

Plasma samples at each timepoint were analyzed using a LC-MS/MS with alower limit of quantification (LLOQ) of 1 ng/mL. Plasma was analyzed todetermine the amount of the compound of the invention in each sample andregression curves generated for each compounds of the invention in therelevant matrixes.

WinNonlin analysis was used for calculating PK parameters for both theIV and PO administrations:

PK parameters for IV portion—AUC_(last); AUC_(INF); T1/2; Cl; Vss;C_(max); MRT

PK parameters for PO portion—AUC_(last); AUC₁; T1/2; C_(max); Cl, MRT.

In addition to the above PK parameters, bioavailability (% F) wascalculated.

Results: Compound 6 had an oral bioavailability of 22%.

Example 25 Efficacy of GSNOR Inhibitors in Experimental InflammatoryBowel Disease (IBD)

Experimental Model

An acute model of dextran sodium sulfate (DSS)-induced IBD in mice isused to explore efficacy of GSNOR inhibitors against this disease. AcuteDSS-induced IBD is a widely used and well characterized model thatinduces pathological changes in the colon similar to those observed inthe human disease. In this model and in human disease, epithelial cellswithin the crypts of the colon are disrupted, leading to dysfunction ofthe epithelial barrier and the ensuing tissue inflammation, edema, andulceration. GSNOR inhibitor therapy may benefit IBD by restorings-nitrosogluthathione (GSNO) levels, and thus prevent or reverse theepithelial barrier dysfunction.

Experimental IBD is induced by administration of DSS in the drinkingwater over several days. GSNOR inhibitors are administered daily viaintravenous (IV) dosing. Effect of treatment is assessed via endoscopyand histopathology using a five point scale ranging from a score=0(normal tissue) to a score=4 (ulcerative tissue damage and markedpathological changes). The effect of GSNOR inhibitors is compared tovehicle treated controls. The corticosteroid, prednisolone, is used asthe positive control in this study and is administered daily via oraldosing. Naïve mice are also assessed as a normal tissue control.

Materials and Methods

Experimental IBD is induced by administration of 3% DSS in the drinkingwater on study days 0 to 5. GSNOR inhibitors are reconstituted toconcentrations of 0.2 and 2 mg/ml in phosphate buffered saline (PBS), pH7.4. Mice are treated daily via IV administration of 0.1 ml GSNORinhibitor solution per mouse for doses of 1 and 10 mg/kg/day. GSNORinhibitor dosing is started 2 days prior to the DSS administration andcontinued through the last day of the study (days −2 to 7). PBS is usedas the vehicle control and is administered in the same manner as theGSNOR inhibitor. The corticosteroid, prednisolone, is used as thepositive control for the study, and is administered orally at a dose of3 mg/kg/day on each day (study days −2 to 7).

The effect of drug treatment is assessed on day 7 via endoscopy andhistopathology. Mice are first anesthetized with inhaled isoflurane andsubjected to endoscopy using a veterinary endoscope (Karl StorzVeterinary Endoscopy America, Inc., Goleta, Calif.). Each mouse isscored for mucosal injury using the endoscopy scoring criteria. Anendoscopy score of 0 is normal, 1 is loss of vascularity, 2 is loss ofvascularity and friability, 3 is friability and erosions, and 4 isulcerations and bleeding. Following endoscopy, mice are euthanized viaasphyxiation with inhaled carbon dioxide. Colon sections are thenformalin-fixed, paraffin-embedded, sectioned, and stained withhematoxylin-eosin. Colon sections are examined via light microscopy andscored in a blinded fashion by a board certified veterinary pathologistwith particular expertise in GI pathology. Pathological changes to theepithelium, connective tissue, and submucosa are scored based oninflammation, edema, and necrosis, and a score of 0 is normal, 1 isminimal, 2 is mild, 3 is moderate, and 4 is marked.

Example 26 Efficacy of GSNOR Inhibitors in Experimental ChronicObstructive Pulmonary Disease (COPD)

Experimental COPD Model

An acute model of elastase-induced COPD in mice is used to exploreefficacy of GSNOR inhibitors against this disease. Elastase-induced COPDis a widely used and well characterized model that induces pathologicalchanges in the lung similar to those observed in the human disease. Inthis model and in human disease, airway obstruction, pulmonaryinflammation, and airspace enlargement are evident. GSNOR inhibitortherapy may benefit COPD through the bronchodilatory andanti-inflammatory actions of these compounds.

Experimental COPD is induced by administration of the elastases, papainand porcine pancreatic elastase (PPE), into the lung over several days.GSNOR inhibitors are administered daily via oral dosing. Efficacy isdetermined by assessing the ability of GSNOR inhibitors to attenuatebronchoconstriction in response to methacholine (MCh) aerosol challenge,decrease pulmonary inflammation, and reduce airspace enlargement in theaveoli. The effect of GSNOR inhibitors are compared to vehicle treatedcontrols. A combination of daily oral SP CXC receptor 2/receptor 1 (SPCXCR2/1) antagonist, which blocks recruitment of neutrophils andmonocytes, and inhaled Flovent (fluticasone; corticosteroid), is used asthe positive control in this study.

Materials and Methods

Experimental COPD is induced by administration of 80 μg papain and 20U/mg PPE per mouse per day via intra-tracheal (IT) instillation on studydays 0 to 7. GSNOR inhibitor is reconstituted to concentrations of 0.01,0.1, and 1 mg/ml in phosphate buffered saline (PBS), pH 7.4. Mice aretreated daily via oral administration (gavage) of 0.1 ml GSNORi solutionper mouse for doses of 0.1, 1, and 10 mg/kg/day. PBS is used as thevehicle control and is administered via daily oral dosing. The smallmolecule antagonist SP CXCR2/R1 (Schering-Plough/Merck), which blocksreceptors to cytokine chemoattractants for neutrophil and monocyterecruitment, is used in combination with the corticosteroid, Flovent(Glaxo), as the positive control for the study. SP CXCR2/R1 is dosedorally at 50 mg/kg/day. Flovent is dosed via inhalation at 220μg/mouse/day. One group of mice is treated with GSNOR inhibitor, vehiclecontrol, or positive control for 7 days (study days 8 to 14), while asecond group of mice is treated with GSNOR inhibitor, vehicle control,or positive control for 14 days (study days 8 to 21).

The effect of drug treatment is assessed 7 and 14 days post-treatment bymeasuring attenuation of methacholine-induced bronchoconstriction(bronchodilatory effect), attenuation of pulmonary inflammation, andreduction of airspace enlargement in the alveoli (14 day post-treatmentonly).

Bronchodilatory Effect

In vivo airway responsiveness to methacholine is measured in conscious,freely moving, spontaneously breathing mice with whole bodyplethysmography using a Buxco chamber (Wilmington, N.C.). Mice arechallenged with aerosolized saline or increasing doses of methacholine(5, 20, and 50 mg/ml) generated by an ultrasonic nebulizer for 2 min.The degree of bronchoconstriction is expressed as enhanced pause (Penh),a calculated dimensionless value, which correlated with the measurementof airway resistance, impedance, and intrapleural pressure in the samemouse. Penh readings are taken and averaged for 4 min. after eachnebulization challenge. Penh is calculated as follows:Penh=[(T_(e)/T_(r)−1)×(PEF/PIF)], where T_(e) is expiration time, T_(r)is relaxation time, PEF is peak expiratory flow, and PIF is peakinspiratory flow×0.67 coefficient. The time for the box pressure tochange from a maximum to a user-defined percentage of the maximumrepresented the relaxation time. The T_(r) measurement began at themaximum box pressure and ended at 40%.

Anti-Inflammatory Effect

After measurement of airway hyper-reactivity, the mice areexsanguination by cardiac puncture, and then bronchoalveolar lavagefluid (BALF) is collected from the right lung after tying off the leftlung at the mainstem bronchus. Total BALF cells are counted, and theremaining fluid is centrifuged at 200×g for 10 min. at 4° C. Cellpellets are resuspended in saline containing 10% bovine serum albumin(BSA) and smears are made on glass slides using cytospin. Cells arestained with Diff-Quik for white blood cell (WBC) differential countsvia light microscopy. Epithelial cells are counted and subtracted fromthe total number of cells. The proportions of eosinophils, macrophages,neutrophils, and lymphocytes are counted using standard morphologicalcriteria and expressed as a percentage of the total number of whiteblood cells (WBCs).

The ability of treatment to reduce levels of neutrophil and monocytechemoattractants in the BALF are also assessed as additional parametersof anti-inflammatory effect. KC (keratinocyte chemoattractant), alsoknown as GROα (growth-related oncogene alpha), and JE (MCP-1, monocytechemoattractant protein), chemokines for neutrophil and monocyterecruitment, respectively, are measured using immunoassay.

Reduction of Airspace Enlargement

Both lungs are inflated under constant positive pressure at 25 cm waterpressure with 10% buffered formaldehyde and then perfused-fixed. Thefixed lungs are embedded in paraffin, stained with hematoxylin andeosin, and examined via light microscopy. Airspace enlargement isquantified morphologically by calculating the mean linear intercept (Lm)and average equivalent diameter of alveoli (D2).

It will be apparent to those skilled in the art that variousmodifications and variations can be made in the methods and compositionsof the present invention without departing from the spirit or scope ofthe invention.

What is claimed is:
 1. A method of treatment of a disease or conditionwherein the disease or condition is selected from the group consistingof pulmonary disorders, inflammatory bowel disease, schizophrenia,psychosis, anxiety, and depression, which comprises administering atherapeutically effective amount of a compound of formula I or apharmaceutically acceptable salt thereof to a patient in need thereof

wherein X is selected from the group consisting of phenyl, substitutedphenyl, thiophen-yl, substituted thiophen-yl, thiazol-yl, substitutedthiazol-yl, pyrazin-yl, substituted pyrazin-yl, pyridin-yl, andsubstituted pyridin-yl, cyclohexyl, substituted cyclohexyl; Y isselected from the group consisting of phenyl, substituted phenyl,thiophen-yl, substituted thiophen-yl, thiazol-yl, substitutedthiazol-yl, pyrazin-yl, substituted pyrazin-yl, pyridin-yl, substitutedpyridin-yl, furan-yl, substituted furan-yl, benzo[d][1,3]dioxol-yl,substituted benzo[d][1,3]dioxol-yl, imidazol-yl, substitutedimidazol-yl, naphthalen-yl, substituted naphthalen-yl, pyrrol-yl,substituted pyrrol-yl, pyrazol-yl, substituted pyrazol-yl,tetrahydrofuran-yl, substituted tetrahydrofuran-yl, cyclopentyl,substituted cyclopentyl, cyclohexyl, and substituted cyclohexyl; Z isselected from the group consisting of 0, S, and NRS; R₁ and R₂ and R₇are independently selected from the group consisting of hydrogen, andC₁-C₆ alkyl; R₃ is selected from the group consisting of hydrogen,nitro, cyano, carboxyl, carbamoyl, methylsulfonamido, fluoro, chloro,bromo, hydroxy, methylsulfonyl, and methylsulfinyl, isoxazol-4-yl, C₁-C₆alkoxy, —C(NH)NHOH, sulfonic acid, and acetyl; R₄ is selected from thegroup consisting of hydroxyl, carboxyl, and tetrazol-5-yl; R₅ isselected from the group consisting of hydrogen, hydroxyl, carboxy,chloro, fluoro, cyano, —O(CH₂)₁₋₆NMe₂, C₁-C₆ alkyl, —O(CH₂)₁₋₆OCH₃,—O(CH₂)₁₋₆OH, acetyl, CF₃, and C₁-C₆ alkoxy; and R₆ is selected from thegroup consisting of hydrogen and hydroxyl.
 2. The method of claim 1wherein R₁, R₂ and R₇ are independently selected from the groupconsisting of hydrogen and methyl; R₅ is selected from the groupconsisting of hydrogen, hydroxyl, carboxyl, chloro, fluoro, cyano,—O(CH₂)₂NMe₂, C₁-C₆ alkyl, —O(CH₂)₂OCH₃, —O(CH₂)₂OH, acetyl, CF₃,methoxy, ethoxy, isopropoxy, and n-propoxy; and R₆ is hydrogen.
 3. Themethod of claim 1 wherein R₃ is selected from the group consisting ofhydrogen, nitro, and hydroxyl; and R₅ is selected from the groupconsisting of hydrogen, ethoxy, fluoro, and —O(CH₂)₂OH.
 4. The method ofclaim 1 wherein X is selected from the group consisting of phenyl,thiophen-2-yl, thiophen-3-yl, thiazol-2-yl, 2-fluorophenyl, p-tolyl,m-tolyl, biphenyl-4-yl, 4-methoxyphenyl, 3-chlorophenyl,3,4-dichlorophenyl, 3-methoxyphenyl, 3,4-dimethoxyphenyl, 4-bromophenyl,o-tolyl, 4-chlorophenyl, 2-chlorophenyl, 3-cyanophenyl,3,4-difluorophenyl, 4-cyanophenyl, 3-carbamoylphenyl, pyrazin-2-yl,biphenyl-3-yl, 2-cyanophenyl, pyridin-4-yl, and pyridin-3-yl,4-(dimethylamino)phenyl, 3-fluorophenyl, 3-ethylphenyl, and cyclohexyl.5. The method of claim 1 wherein X is selected from the group consistingof phenyl, thiophen-2-yl, thiophen-3-yl, and pyridin-3-yl.
 6. The methodof claim 1 wherein Y is selected from the group consisting of phenyl,3-methoxyphenyl, p-tolyl, 4-methoxyphenyl, 3,5-dichlorophenyl,3-fluorophenyl, 4-bromophenyl, biphenyl-4-yl, 4-fluorophenyl,4-chlorophenyl, 3-chlorophenyl, 3,4-dimethoxyphenyl,3-fluoro-4-methoxyphenyl, 4-chloro-3-fluorophenyl,3-chloro-4-fluorophenyl, 3,4-difluorophenyl, 3,5-difluorophenyl,3,4-dichlorophenyl, 4-hydroxyphenyl, 2,4-difluorophenyl, furan-3-yl,2-chlorophenyl, 3-cyanophenyl, 4-(dimethylamino)phenyl, 2-fluorophenyl,4-morpholinophenyl, 4-aminophenyl, pyridin-2-yl,benzo[d][1,3]dioxol-5-yl, 4-cyanophenyl, pyridin-3-yl, pyridin-4-yl,4-acetamidophenyl, thiophen-2-yl, thiophen-3-yl,1-methyl-1H-imidazol-4-yl, naphthalen-1-yl, methyl phenylcarbamate, andnaphthalen-2-yl, 4-(methanesulfonamido)phenyl, 1H-pyrrol-3-yl,1-(phenylsulfonyl)-1H-pyrrol-3-yl, furan-2-yl,4-(trifluoromethyl)phenyl, o-tolyl, 1-methyl-1H-pyrazol-4-yl,1-methyl-1H-pyrazol-3-yl, 3-chloro-5-fluorophenyl, 3-hydroxyphenyl,pyrazin-2-yl, quinolin-6-yl, isoquinolin-6-yl, 1-methyl-1H-pyrazol-5-yl,tetrahydrofuran-2-yl, cyclopentyl, tetrahydrofuran-3-yl and cyclohexyl.7. The method of claim 1 wherein Y is selected from the group consistingof phenyl, pyridin-3-yl, 1-methyl-1H-pyrazol-4-yl, and cyclohexyl. 8.The method of claim 1 wherein Z is O.
 9. The method of claim 1 whereinthe compound is selected from the group consisting of4-(3-ethoxy-4-hydroxy-5-nitrophenyl)-5,6-diphenyl-3,4-dihydropyridin-2(1H)-one;4-(3-ethoxy-4-hydroxy-5-nitrophenyl)-6-(pyridin-3-yl)-5-(thiophen-3-yl)-3,4-dihydropyridin-2(1H)-one;4-(4-(2H-tetrazol-5-yl)phenyl)-5-phenyl-6-(pyridin-3-yl)-3,4-dihydropyridin-2(1H)-one;4-(3-ethoxy-4-hydroxy-5-nitrophenyl)-6-(1-methyl-1H-pyrazol-4-yl)-5-(thiophen-3-yl)-3,4-dihydropyridin-2(1H)-one;4-(4-(2H-tetrazol-5-yl)phenyl)-6-(1-methyl-1H-pyrazol-4-yl)-5-(thiophen-3-yl)-3,4-dihydropyridin-2(1H)-one;2-hydroxy-4-(2-oxo-5,6-diphenyl-1,2,3,4-tetrahydropyridin-4-yl)benzoicacid;2-hydroxy-4-(2-oxo-6-phenyl-5-(thiophen-3-yl)-1,2,3,4-tetrahydropyridin-4-yl)benzoicacid;4-(4-(2H-tetrazol-5-yl)phenyl)-6-(1-methyl-1H-pyrazol-4-yl)-5-phenyl-3,4-dihydropyridin-2(1H)-one;2-hydroxy-4-(6-(1-methyl-1H-pyrazol-4-yl)-2-oxo-5-(thiophen-3-yl)-1,2,3,4-tetrahydropyridin-4-yl)benzoicacid;2-fluoro-6-hydroxy-4-(2-oxo-5,6-diphenyl-1,2,3,4-tetrahydropyridin-4-yl)benzoicacid;2-fluoro-6-hydroxy-4-(6-(1-methyl-1H-pyrazol-4-yl)-2-oxo-5-(thiophen-3-yl)-1,2,3,4-tetrahydropyridin-4-yl)benzoicacid;2-ethoxy-6-hydroxy-4-(2-oxo-5,6-diphenyl-1,2,3,4-tetrahydropyridin-4-yl)benzoicacid;4-(6-cyclohexyl-2-oxo-5-(thiophen-3-yl)-1,2,3,4-tetrahydropyridin-4-yl)-2-fluoro-6-hydroxybenzoicacid;4-(4-hydroxy-3-(2-hydroxyethoxy)-5-nitrophenyl)-5,6-diphenyl-3,4-dihydropyridin-2(1H)-one;2-fluoro-6-hydroxy-4-(1-methyl-2-oxo-5,6-diphenyl-1,2,3,4-tetrahydropyridin-4-yl)benzoicacid;4-(3-ethoxy-4-hydroxy-5-nitrophenyl)-1-methyl-5,6-diphenyl-3,4-dihydropyridin-2(1H)-one;and2-fluoro-6-hydroxy-4-(6-(1-methyl-1H-pyrazol-4-yl)-2-oxo-5-(thiophen-2-yl)-1,2,3,4-tetrahydropyridin-4-yl)benzoicacid.
 10. The method of claim 1 wherein the disease or condition is apulmonary disorder.
 11. The method of claim 10 wherein the pulmonarydisorder is selected from the group consisting of asthma, cysticfibrosis, and chronic obstructive pulmonary disorder (COPD).
 12. Themethod of claim 1 wherein the disease or condition is inflammatory boweldisease (IBD).
 13. The method of claim 1 wherein the disease orcondition is selected from the group consisting of anxiety, depression,psychosis and schizophrenia.
 14. A method of inhibitingS-nitrosoglutathione reductase (GSNOR) in a subject, said methodcomprising administering to the subject a pharmacologically effectiveamount of a pharmaceutical composition comprising the compound offormula I as defined in claim 1 or a pharmaceutically acceptable saltthereof.
 15. The method of claim 14 wherein the compound of formula I isthe S enantiomer.
 16. The method of claim 14 wherein the compound isselected from the group consisting of4-(3-ethoxy-4-hydroxy-5-nitrophenyl)-5,6-diphenyl-3,4-dihydropyridin-2(1H)-one;4-(3-ethoxy-4-hydroxy-5-nitrophenyl)-6-(pyridin-3-yl)-5-(thiophen-3-yl)-3,4-dihydropyridin-2(1H)-one;4-(4-(2H-tetrazol-5-yl)phenyl)-5-phenyl-6-(pyridin-3-yl)-3,4-dihydropyridin-2(1H)-one;4-(3-ethoxy-4-hydroxy-5-nitrophenyl)-6-(1-methyl-1H-pyrazol-4-yl)-5-(thiophen-3-yl)-3,4-dihydropyridin-2(1H)-one;4-(4-(2H-tetrazol-5-yl)phenyl)-6-(1-methyl-1H-pyrazol-4-yl)-5-(thiophen-3-yl)-3,4-dihydropyridin-2(1H)-one;2-hydroxy-4-(2-oxo-5,6-diphenyl-1,2,3,4-tetrahydropyridin-4-yl)benzoicacid;2-hydroxy-4-(2-oxo-6-phenyl-5-(thiophen-3-yl)-1,2,3,4-tetrahydropyridin-4-yl)benzoicacid;4-(4-(2H-tetrazol-5-yl)phenyl)-6-(1-methyl-1H-pyrazol-4-yl)-5-phenyl-3,4-dihydropyridin-2(1H)-one;2-hydroxy-4-(6-(1-methyl-1H-pyrazol-4-yl)-2-oxo-5-(thiophen-3-yl)-1,2,3,4-tetrahydropyridin-4-yl)benzoicacid;2-fluoro-6-hydroxy-4-(2-oxo-5,6-diphenyl-1,2,3,4-tetrahydropyridin-4-yl)benzoicacid;2-fluoro-6-hydroxy-4-(6-(1-methyl-1H-pyrazol-4-yl)-2-oxo-5-(thiophen-3-yl)-1,2,3,4-tetrahydropyridin-4-yl)benzoicacid;2-ethoxy-6-hydroxy-4-(2-oxo-5,6-diphenyl-1,2,3,4-tetrahydropyridin-4-yl)benzoicacid;4-(6-cyclohexyl-2-oxo-5-(thiophen-3-yl)-1,2,3,4-tetrahydropyridin-4-yl)-2-fluoro-6-hydroxybenzoicacid;4-(4-hydroxy-3-(2-hydroxyethoxy)-5-nitrophenyl)-5,6-diphenyl-3,4-dihydropyridin-2(1H)-one;2-fluoro-6-hydroxy-4-(1-methyl-2-oxo-5,6-diphenyl-1,2,3,4-tetrahydropyridin-4-yl)benzoicacid;4-(3-ethoxy-4-hydroxy-5-nitrophenyl)-1-methyl-5,6-diphenyl-3,4-dihydropyridin-2(1H)-one;and2-fluoro-6-hydroxy-4-(6-(1-methyl-1H-pyrazol-4-yl)-2-oxo-5-(thiophen-2-yl)-1,2,3,4-tetrahydropyridin-4-yl)benzoicacid.
 17. A method of antagonizing the tachkinin receptor NK3 in asubject, said method comprising administering to the subject apharmacologically effective amount of a pharmaceutical compositioncomprising the compound of formula I as defined in claim 1 or apharmaceutically acceptable salt thereof.
 18. The method of claim 17wherein the compound of formula I is the R enantiomer.
 19. The method ofclaim 17 wherein the compound is selected from the group consisting of4-(3-ethoxy-4-hydroxy-5-nitrophenyl)-5,6-diphenyl-3,4-dihydropyridin-2(1H)-one;4-(3-ethoxy-4-hydroxy-5-nitrophenyl)-6-(pyridin-3-yl)-5-(thiophen-3-yl)-3,4-dihydropyridin-2(1H)-one4-(3-ethoxy-4-hydroxy-5-nitrophenyl)-6-(1-methyl-1H-pyrazol-4-yl)-5-(thiophen-3-yl)-3,4-dihydropyridin-2(1H)-one;4-(4-hydroxy-3-(2-hydroxyethoxy)-5-nitrophenyl)-5,6-diphenyl-3,4-dihydropyridin-2(1H)-one;and4-(3-ethoxy-4-hydroxy-5-nitrophenyl)-1-methyl-5,6-diphenyl-3,4-dihydropyridin-2(1H)-one.